INHOUD
Inleiding................................................................................................................................................................... 5
H2: Biomoleculen .................................................................................................................................................... 5
2.1 Bio-elementen en bio-moleculen .................................................................................................................. 5
2.2 Water ............................................................................................................................................................ 6
2.3 Andere atomen en moleculen ....................................................................................................................... 7
C,O,N, H ........................................................................................................................................................... 7
Anorganische stoffen ...................................................................................................................................... 7
Fosfor .............................................................................................................................................................. 8
Zwavel ............................................................................................................................................................. 9
Jood ................................................................................................................................................................. 9
ijzer ................................................................................................................................................................ 10
Organische molc ............................................................................................................................................ 11
Gluciden ........................................................................................................................................................ 11
Aldose, ketosen ......................................................................................................................................... 13
Disacariden ................................................................................................................................................ 14
Polysachariden .......................................................................................................................................... 14
Glycoproteïnen en glycolipiden (O en N gebonden) ................................................................................. 15
Aanverwante moleculen ........................................................................................................................... 16
Lipiden ........................................................................................................................................................... 18
2.4 In stand houden vd homeostasis ................................................................................................................ 21
H3: pH en buffermensels ...................................................................................................................................... 22
3.1 Ionisatie van water ...................................................................................................................................... 22
3.2 Zuurtegraad of pH ....................................................................................................................................... 22
3.3 pH van zuren en basen ................................................................................................................................ 22
3.4 pH buffermengsel ........................................................................................................................................ 22
3.5 pH in het menselijk organisme .................................................................................................................... 22
3.5.1 Buffersystemen in het bloed ................................................................................................................ 22
H2CO3 en NaHCO3- systeem .................................................................................................................... 22
Fosfaat buffersysteem............................................................................................................................... 23
Hemoglobine buffersysteem ..................................................................................................................... 23
3.5.2 Longfunctie........................................................................................................................................... 23
H2CO3 ....................................................................................................................................................... 23
H2SO4 en H3PO4....................................................................................................................................... 23
3.5.3 NIerfunctie ........................................................................................................................................... 24
3.5.4 Factoren die invloed hebben op de pH van het bloed ......................................................................... 24
, 3.5.5 Acidose en alkalose .............................................................................................................................. 25
acidose ...................................................................................................................................................... 25
Alkalose ..................................................................................................................................................... 25
H4: Biochemische technieken ............................................................................................................................... 29
4.1 Elektroforese ............................................................................................................................................... 29
Soorten dragers ............................................................................................................................................ 29
Cellulose of papier..................................................................................................................................... 30
Celluloseacetaat ........................................................................................................................................ 30
Agar ........................................................................................................................................................... 30
Agarose ..................................................................................................................................................... 30
polyacrylamide .......................................................................................................................................... 31
Capillair ..................................................................................................................................................... 31
Matrix/ drager .............................................................................................................................................. 31
Elektrische mobiliteit ion ............................................................................................................................. 31
Aangelegde spanning ................................................................................................................................ 31
Netto lading deeltje .................................................................................................................................. 32
Wrijving deeltjes ....................................................................................................................................... 32
belang pH vd buffer ...................................................................................................................................... 32
kleuring ......................................................................................................................................................... 33
Densitometrie ............................................................................................................................................... 34
4.1.1 Cellulose acetaat elektroforese................................................................................................................ 34
4.1.2 Polyacrylamide gelelektroforese (PAGE) voor EW ................................................................................... 36
gel ( polyacrylamide gel ) .............................................................................................................................. 36
Continue en discontinue buffersystemen ..................................................................................................... 37
SDS-PAGE de denaturerende gelelektroforese ............................................................................................. 39
4.1.3 Agarose gelelektroforese ......................................................................................................................... 41
4.1.4 Immunofixatie .......................................................................................................................................... 42
4.1.5 Iso- elektrische focussering ( IEF) ............................................................................................................. 42
Toepassingen ................................................................................................................................................ 44
Opsporen multiple sclerose in cerebrospinaal vocht ................................................................................ 44
Bepaling HbA1c ......................................................................................................................................... 44
opsporen EPO ( erytropoëtine) in urine ........................................................................................................ 44
4.1.6 2D Gelelektroforese ................................................................................................................................. 46
4.1.7 Capillaire elektroforese (CE) ..................................................................................................................... 47
4.2 Immunoblotting .......................................................................................................................................... 49
4.3 Kolomchromatografie ................................................................................................................................. 51
gelfiltratie ...................................................................................................................................................... 51
, Ionwisselingschromatografie ........................................................................................................................ 51
Affiniteitschromatografie .............................................................................................................................. 53
Toepassingen affiniteitschromatografie ................................................................................................... 53
Vergelijking met page................................................................................................................................ 54
4.4 Uit en inzouten van EW ............................................................................................................................... 55
4.5 Dialyse ......................................................................................................................................................... 56
4.6 Centrifugering ............................................................................................................................................. 56
4.7 Celfractionering ........................................................................................................................................... 59
H.5 Proteïnen ........................................................................................................................................................ 61
5.1 Inleiding ....................................................................................................................................................... 61
5.2 Aminozuren ................................................................................................................................................ 63
Notities en ppt .............................................................................................................................................. 63
UIT CURSUS ................................................................................................................................................... 66
5.3 Eiwitten ....................................................................................................................................................... 67
5.4 sTRUCTUUR EIWITTEN ............................................................................................................................... 78
ALFA-KERATINE ............................................................................................................................................. 78
Collageen ....................................................................................................................................................... 78
Zijdefibroïne .................................................................................................................................................. 79
5.5 Globulaire Eiwitten ...................................................................................................................................... 79
Eiwitdynamiek ............................................................................................................................................... 80
5.6 vouwen van eiwitten ................................................................................................................................... 80
vouwpatronen ............................................................................................................................................... 80
Biologisch actieve conformatie ..................................................................................................................... 80
Foutief gevouwen eiwitten (inactief of gevaarlijk) ....................................................................................... 80
Denatureren en renatureren ........................................................................................................................ 81
Interacties van diverse producten ................................................................................................................ 81
7.2 Voorbeelden ................................................................................................................................................ 83
5.9 Sequentiebepaling en evolutie van eiwitten ............................................................................................... 86
5.10 Eiwitten in het labo ................................................................................................................................... 87
Eiwit precipitatietechniek/onteiwitten ......................................................................................................... 87
Assay voor eiwitbepaling .............................................................................................................................. 87
Analyse voor identificatie en structuur ......................................................................................................... 87
H6 Enzymen en coënzymen .................................................................................................................................. 88
6.1 Inleiding ....................................................................................................................................................... 88
6.2 Mechanisme v. katalyse .............................................................................................................................. 88
6.3 Eigenschappen van enzymen ...................................................................................................................... 89
6.4 Coënzymen .................................................................................................................................................. 89
, 6.5 Enzymkinetiek ............................................................................................................................................. 91
6.6 Indeling van enzymes .................................................................................................................................. 93
6.7 Vitamen ....................................................................................................................................................... 95
H7: Overzicht van het metabolisme ...................................................................................................................... 95
1 Anabolisme en katabolisme ........................................................................................................................... 95
2. Energiecycli in de cel: 1 de ATP/ADP cyclus .................................................................................................. 96
2: Energiecycli in de cel: 2 de NAD+/NADH + H+ cyclus .................................................................................... 97
H8: Metabolisme van koolhydraten ..................................................................................................................... 98
1. Inleiding ......................................................................................................................................................... 98
2 Glycolyse ........................................................................................................................................................ 98
2.1 Verband tss fermentatie en ademhaling................................................................................................. 98
2.2 overzicht glocolyse: zie vanaf p 11 ........................................................................................................ 100
3. Pentose cyclus of “hexose-monofosfaat shunt” ......................................................................................... 100
4. Afbraak en opbouw van glycogeen ............................................................................................................. 100
4.1 Afbraak van glycogeen tot glycogeneolyse ............................................................................................... 101
4.2 Opbouw van glycogeen of glycogenogenese ............................................................................................ 101
9. Biochemische energie ..................................................................................................................................... 102
9.2 Energie-rijke verbindingen ........................................................................................................................ 102
9.3Citroenzuur-of krebcyclus .......................................................................................................................... 102
9.3.1 Metabolisme v pyrodruivenzuur ........................................................................................................ 102
9.3.2 Overzicht van de Kreb cyclus = citroenzuurcyclus ............................................................................. 102
9.3.3 Individuele reacties kreb-cyclus ........................................................................................................ 103
9.4 Elektronentransport en oxidatieve fosfoylatie ......................................................................................... 104
9.5 Schatting energieopbrengst bij oxidatie glucose ...................................................................................... 105
10 Metabolisme van proteïnen ........................................................................................................................... 106
11 Metabolisme van lipiden ............................................................................................................................... 106
Opbouw en afbraak v lipiden .......................................................................................................................... 106
H12: Signaaltransductie ...................................................................................................................................... 109
https://www.youtube.com/watch?v=LHn3p2bjO7U
Inleiding................................................................................................................................................................... 5
H2: Biomoleculen .................................................................................................................................................... 5
2.1 Bio-elementen en bio-moleculen .................................................................................................................. 5
2.2 Water ............................................................................................................................................................ 6
2.3 Andere atomen en moleculen ....................................................................................................................... 7
C,O,N, H ........................................................................................................................................................... 7
Anorganische stoffen ...................................................................................................................................... 7
Fosfor .............................................................................................................................................................. 8
Zwavel ............................................................................................................................................................. 9
Jood ................................................................................................................................................................. 9
ijzer ................................................................................................................................................................ 10
Organische molc ............................................................................................................................................ 11
Gluciden ........................................................................................................................................................ 11
Aldose, ketosen ......................................................................................................................................... 13
Disacariden ................................................................................................................................................ 14
Polysachariden .......................................................................................................................................... 14
Glycoproteïnen en glycolipiden (O en N gebonden) ................................................................................. 15
Aanverwante moleculen ........................................................................................................................... 16
Lipiden ........................................................................................................................................................... 18
2.4 In stand houden vd homeostasis ................................................................................................................ 21
H3: pH en buffermensels ...................................................................................................................................... 22
3.1 Ionisatie van water ...................................................................................................................................... 22
3.2 Zuurtegraad of pH ....................................................................................................................................... 22
3.3 pH van zuren en basen ................................................................................................................................ 22
3.4 pH buffermengsel ........................................................................................................................................ 22
3.5 pH in het menselijk organisme .................................................................................................................... 22
3.5.1 Buffersystemen in het bloed ................................................................................................................ 22
H2CO3 en NaHCO3- systeem .................................................................................................................... 22
Fosfaat buffersysteem............................................................................................................................... 23
Hemoglobine buffersysteem ..................................................................................................................... 23
3.5.2 Longfunctie........................................................................................................................................... 23
H2CO3 ....................................................................................................................................................... 23
H2SO4 en H3PO4....................................................................................................................................... 23
3.5.3 NIerfunctie ........................................................................................................................................... 24
3.5.4 Factoren die invloed hebben op de pH van het bloed ......................................................................... 24
, 3.5.5 Acidose en alkalose .............................................................................................................................. 25
acidose ...................................................................................................................................................... 25
Alkalose ..................................................................................................................................................... 25
H4: Biochemische technieken ............................................................................................................................... 29
4.1 Elektroforese ............................................................................................................................................... 29
Soorten dragers ............................................................................................................................................ 29
Cellulose of papier..................................................................................................................................... 30
Celluloseacetaat ........................................................................................................................................ 30
Agar ........................................................................................................................................................... 30
Agarose ..................................................................................................................................................... 30
polyacrylamide .......................................................................................................................................... 31
Capillair ..................................................................................................................................................... 31
Matrix/ drager .............................................................................................................................................. 31
Elektrische mobiliteit ion ............................................................................................................................. 31
Aangelegde spanning ................................................................................................................................ 31
Netto lading deeltje .................................................................................................................................. 32
Wrijving deeltjes ....................................................................................................................................... 32
belang pH vd buffer ...................................................................................................................................... 32
kleuring ......................................................................................................................................................... 33
Densitometrie ............................................................................................................................................... 34
4.1.1 Cellulose acetaat elektroforese................................................................................................................ 34
4.1.2 Polyacrylamide gelelektroforese (PAGE) voor EW ................................................................................... 36
gel ( polyacrylamide gel ) .............................................................................................................................. 36
Continue en discontinue buffersystemen ..................................................................................................... 37
SDS-PAGE de denaturerende gelelektroforese ............................................................................................. 39
4.1.3 Agarose gelelektroforese ......................................................................................................................... 41
4.1.4 Immunofixatie .......................................................................................................................................... 42
4.1.5 Iso- elektrische focussering ( IEF) ............................................................................................................. 42
Toepassingen ................................................................................................................................................ 44
Opsporen multiple sclerose in cerebrospinaal vocht ................................................................................ 44
Bepaling HbA1c ......................................................................................................................................... 44
opsporen EPO ( erytropoëtine) in urine ........................................................................................................ 44
4.1.6 2D Gelelektroforese ................................................................................................................................. 46
4.1.7 Capillaire elektroforese (CE) ..................................................................................................................... 47
4.2 Immunoblotting .......................................................................................................................................... 49
4.3 Kolomchromatografie ................................................................................................................................. 51
gelfiltratie ...................................................................................................................................................... 51
, Ionwisselingschromatografie ........................................................................................................................ 51
Affiniteitschromatografie .............................................................................................................................. 53
Toepassingen affiniteitschromatografie ................................................................................................... 53
Vergelijking met page................................................................................................................................ 54
4.4 Uit en inzouten van EW ............................................................................................................................... 55
4.5 Dialyse ......................................................................................................................................................... 56
4.6 Centrifugering ............................................................................................................................................. 56
4.7 Celfractionering ........................................................................................................................................... 59
H.5 Proteïnen ........................................................................................................................................................ 61
5.1 Inleiding ....................................................................................................................................................... 61
5.2 Aminozuren ................................................................................................................................................ 63
Notities en ppt .............................................................................................................................................. 63
UIT CURSUS ................................................................................................................................................... 66
5.3 Eiwitten ....................................................................................................................................................... 67
5.4 sTRUCTUUR EIWITTEN ............................................................................................................................... 78
ALFA-KERATINE ............................................................................................................................................. 78
Collageen ....................................................................................................................................................... 78
Zijdefibroïne .................................................................................................................................................. 79
5.5 Globulaire Eiwitten ...................................................................................................................................... 79
Eiwitdynamiek ............................................................................................................................................... 80
5.6 vouwen van eiwitten ................................................................................................................................... 80
vouwpatronen ............................................................................................................................................... 80
Biologisch actieve conformatie ..................................................................................................................... 80
Foutief gevouwen eiwitten (inactief of gevaarlijk) ....................................................................................... 80
Denatureren en renatureren ........................................................................................................................ 81
Interacties van diverse producten ................................................................................................................ 81
7.2 Voorbeelden ................................................................................................................................................ 83
5.9 Sequentiebepaling en evolutie van eiwitten ............................................................................................... 86
5.10 Eiwitten in het labo ................................................................................................................................... 87
Eiwit precipitatietechniek/onteiwitten ......................................................................................................... 87
Assay voor eiwitbepaling .............................................................................................................................. 87
Analyse voor identificatie en structuur ......................................................................................................... 87
H6 Enzymen en coënzymen .................................................................................................................................. 88
6.1 Inleiding ....................................................................................................................................................... 88
6.2 Mechanisme v. katalyse .............................................................................................................................. 88
6.3 Eigenschappen van enzymen ...................................................................................................................... 89
6.4 Coënzymen .................................................................................................................................................. 89
, 6.5 Enzymkinetiek ............................................................................................................................................. 91
6.6 Indeling van enzymes .................................................................................................................................. 93
6.7 Vitamen ....................................................................................................................................................... 95
H7: Overzicht van het metabolisme ...................................................................................................................... 95
1 Anabolisme en katabolisme ........................................................................................................................... 95
2. Energiecycli in de cel: 1 de ATP/ADP cyclus .................................................................................................. 96
2: Energiecycli in de cel: 2 de NAD+/NADH + H+ cyclus .................................................................................... 97
H8: Metabolisme van koolhydraten ..................................................................................................................... 98
1. Inleiding ......................................................................................................................................................... 98
2 Glycolyse ........................................................................................................................................................ 98
2.1 Verband tss fermentatie en ademhaling................................................................................................. 98
2.2 overzicht glocolyse: zie vanaf p 11 ........................................................................................................ 100
3. Pentose cyclus of “hexose-monofosfaat shunt” ......................................................................................... 100
4. Afbraak en opbouw van glycogeen ............................................................................................................. 100
4.1 Afbraak van glycogeen tot glycogeneolyse ............................................................................................... 101
4.2 Opbouw van glycogeen of glycogenogenese ............................................................................................ 101
9. Biochemische energie ..................................................................................................................................... 102
9.2 Energie-rijke verbindingen ........................................................................................................................ 102
9.3Citroenzuur-of krebcyclus .......................................................................................................................... 102
9.3.1 Metabolisme v pyrodruivenzuur ........................................................................................................ 102
9.3.2 Overzicht van de Kreb cyclus = citroenzuurcyclus ............................................................................. 102
9.3.3 Individuele reacties kreb-cyclus ........................................................................................................ 103
9.4 Elektronentransport en oxidatieve fosfoylatie ......................................................................................... 104
9.5 Schatting energieopbrengst bij oxidatie glucose ...................................................................................... 105
10 Metabolisme van proteïnen ........................................................................................................................... 106
11 Metabolisme van lipiden ............................................................................................................................... 106
Opbouw en afbraak v lipiden .......................................................................................................................... 106
H12: Signaaltransductie ...................................................................................................................................... 109
https://www.youtube.com/watch?v=LHn3p2bjO7U
