Enzyme Mechanisms

BIOC12

Covalent catalysis

covalent bond between substrate and enzyme creates unstable intermediate which promotes catalysis

Acid Catalysis

donation of proton by enzyme

Base catalysis

removal of proton by enzyme

Metal-ion catalysis

metal ions act as cofactors that promote orientation of bound substrates or shield/stabilize charges. can also create nucleophile by increasing acidity of nearby molecules

metal-activiated enzymes

loosely bound enzymes, doesn’t need metal ions but are helpful

metalloenzymes

tightly bound enzymes, metal ion required

Proximity effect

reactions with two distinct substrates could have enhanced rate by being in close proximity or proper orientation

Chymotrypsin

Serine protease that selectively cleaves peptide bonds on carboxyl side of Trp, Tyr, Phe, Met, Ile

Catalytic Triad

3 AAs that create H-bonded network for catalysis (Ser195, His57, Asp102)

Ser195

Catalytic residue that forms ES covalent intermediate

His57

General base catalyst: accepts H from Ser195, making it a reactive nucleophile

Asp102

Orient His57 and improve its basic properties

Chymotrypsin Phase 1

Formation of a covalent enzyme intermediate between Ser195 and substrate (promotes cleavage of peptide bond and release of C-terminal fragment)

Chymotrypsin Phase 2

Enzyme regeneration based on deacylation and release of N-terminal fragment

Enolase

Metalloenzyme that catalyzes dehydration of 2-phosphoglycerate to PEP, contains 2 divalent metal ions (Mg2+)

Enolase Catalytic Strategies

Lys435 as a general base and Glu211 as general acid

HMG-CoA Reductase

Enzyme for cholesterol, important drug target for hypercholesterolemia

Inhibition of HMG-CoA

Reduction in cholesterol production, lowering serum cholesterol and risk of heart disease

Coenzyme-dependent redox reaction

Transfer of electrons between coenzyme and substrate so one is oxidized and other is reduced

Metabolite transformation reactions

Chemical transformation of metabolites to reactive intermediates (needed for ana/catabolic reactions)

Reversible covalent modification reactions

Attachment/removal of molecular tags to control activity, such as cell signalling or gene expression

Phosphorylation targets

Ser, Thr, Tyr

Adenylation

Addition of nucleoside monophosphate

Zymogens

Enzyme inactive forms that can be irreversibly activated by cleavage, for transfer

Chymotrypsin step 1

Enzyme binding substrate (aromatic side chain, specificity pocket lining)

Chymotrypsin step 2

H transfer from Ser195 to His57 = Ser195 is a nucleophile that attacks C=O carbon on peptide backbone, formation of tetrahedral intermediate with oxyanion that H-bonds with NH of Ser195 and Gly193

Chymotrypsin step 3

Imidazole of His57 donates H to N of peptide bond = cleavage and release of C-terminal fragment, N-terminal fragment bound as acyl-enzyme intermediate

Chymotrypsin step 4

Water donates H To His = free OH that attacks C=O carbon on acyl-enzyme = second tetrahedral intermediate (stabilized by oxyanion hole)

Chymotrypsin step 5

Protonated His57 donates H to cleave covalent bond of acyl-enzyme intermediate, N-terminal fragment released

Chymotrypsin step 6

Catalytic triad regenerated

HMG-CoA Reductase Mechanism

NADPH induces production of mevalonate from HMG-CoA through H transfers with Glu

NAD typically involve ? bonds

C-O

FAD typically involve ? bonds

C-C