Enzyme mechanism Flashcards

1
Q

if 2S and 2P

A

A+B C+D
M-M kinetic - different set up
measure 4 different Km

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2
Q

change [S] and measure Rate

A

each keep S - saturated level and change other conc

e.g. Km for A - saturated [B] and gradual increase of [A]

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3
Q

multiple S/P reaction

A

sequential method

ping pong method

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4
Q

sequential method

A

E+A+B -> EAB -> ECD -> E+C+D

binding might be ordered (bind first and other bind next) or random

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5
Q

ping pong method

A

A+B never meet and E at same time
E+A -> EA -> E.C -> Eā€™ -> EB -> ED -> E+D
Eā€™ - modified

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6
Q

activation energy

A

change in energy to small multiple step using E
E+S -> ES -> ET -> EP -> E+P
binding energy partially offsets Ae

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7
Q

E strategies

A

general

specific chemical

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8
Q

general E strategies

A

position of R
distortion of R
stabilisation of ET
provision enclosed chemical environment

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9
Q

general - position of R

A

position R correctly for interaction - bind to each other - one angle

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10
Q

general - distortion of R

A

less stable therefore undergo reactivity

change in environment - favour reaction

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11
Q

general - stabilisation of ET

A

affinity -> ET>S>P

bind and orders of magnitude more strongly than S

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12
Q

general - provision

A

enclosed chemical environment

e.g. pH differ from outside

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13
Q

specific chemical

A

3 catalysis: covalent, acid/base and metal ion

combo of non-covalent binding effect and covalent chemical interaction

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14
Q

covalent catalysis

A

a.s.r react with S

modify bond and change energy disruption

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15
Q

a.s.r

A

active site residue

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16
Q

acid/base catalysis

A

a.s.r accept/donate H+
basic and acidic a.a
Cys, Ser, Tyr

17
Q

metal ion catalysis

A

various type

18
Q

chemical strategies - proteolysis

A

hydrolyse peptide bond by boiling in 6M HCl for 24hrs

19
Q

serine protease

A

Gene duplication and divergence

same reaction but different S

20
Q

example serine protease

A

trypsin
chymotrypsin
elastase (all found in digestive juice)
thrombin (blood clotting)

21
Q

specificity by pocket

A

region in enzyme - gives specificity
e.g. chymotrypsin, trypsin, elastase
main polypeptide line up and cut at same relative position

22
Q

specificity by pocket - chymotrypsin

A

empty space for large a.a

23
Q

specificity by pocket - trypsin

A

-ve bind strongly to +ve charge S

24
Q

specificity by pocket - elastase

A

bulky HP side chain = narrow space - small side chain

25
Q

chymotrypsin

A

catalytic triad (D/H/S) - positioned - Ser-His-Asp

26
Q

mechanism - chymotrypsin (1)

A

Nu- attach on polypeptide carbonyl-acid/base catalysis -> alkoxide ion form bond with C on S chain

27
Q

mechanism - chymotrypsin (2)

A

covalent intermediate -> S covalent attach to E = tetrahedral intermediate = destabilisation of S - oxyanion hole

28
Q

mechanism - chymotrypsin (3)

A

cleave and lose C-terminal fragment -> leaves as peptide bond broken

29
Q

mechanism - chymotrypsin (4)

A

Nu- attach on polypeptide carbonyl by H2O - H removed by His = OH- Nu- - acid/base catalysis

30
Q

mechanism - chymotrypsin (5)

A

covalent intermediate - regenerate carbonyl at N term is = tetrahedral = set retrieve H from N of His and covalent bond with Ser - 0

31
Q

mechanism - chymotrypsin (6)

A

cleave and loss of N-terminal fragment - OG position

32
Q

oxyanion hole

A

always present inactive site
shown 0 can move in when single bonded
planar to tetrahedral -> 0 form H with NH of gly109 and Ser195

33
Q

example oxyanion hole

A

cysteine protease - H lost at Cys = sulphur Nu-

metalloprotease - Nu- attach Zinc

34
Q

DIDF

A

react with Ser195

35
Q

TPCK

A

react with His57