Specificity in deacylation of acyl-α-chymotrypsins

Ingles, D. W.; Knowles, Jeremy R.; Tomlinson, James

doi:10.1016/0006-291x(66)90444-x

Cited by 10 publications

(4 citation statements)

References 11 publications

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“…Incubation of enzyme-inhibitor complex with hydroxylamine restored 70%–80% activity of GlpG within 30 min (Figure S4C). Analogous to chymotrypsin, where the rate of deacylation was observed to be dependent on the chemical groups (Ingles et al., 1966), the difference in the rate of deacylation of β-lactams in GlpG may reflect the nature of each hydrophobic group’s interaction with the enzyme. Relatively slow deacylation of β-lactams indicates that they are poor substrates for GlpG, forming a nonproductive structure and explains why a stable acyl enzyme complex could be observed in the crystals.…”

Section: Resultsmentioning

confidence: 99%

Structure of Rhomboid Protease in Complex with β-Lactam Inhibitors Defines the S2′ Cavity

et al. 2013

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SummaryRhomboids are evolutionarily conserved serine proteases that cleave transmembrane proteins within the membrane. The increasing number of known rhomboid functions in prokaryotes and eukaryotes makes them attractive drug targets. Here, we describe structures of the Escherichia coli rhomboid GlpG in complex with β-lactam inhibitors. The inhibitors form a single bond to the catalytic serine and the carbonyl oxygen of the inhibitor faces away from the oxyanion hole. The hydrophobic N-substituent of β-lactam inhibitors points into a cavity within the enzyme, providing a structural explanation for the specificity of β-lactams on rhomboid proteases. This same cavity probably represents the S2′ substrate binding site of GlpG. We suggest that the structural changes in β-lactam inhibitor binding reflect the state of the enzyme at an initial stage of substrate binding to the active site. The structural insights from these enzyme-inhibitor complexes provide a starting point for structure-based design for rhomboid inhibitors.

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Section: Resultsmentioning

confidence: 99%

Structure of Rhomboid Protease in Complex with β-Lactam Inhibitors Defines the S2′ Cavity

et al. 2013

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“…This is in agreement with the finding of Deranleau & Neurath (1966) that the binding to oc-chymotrypsin of the inhibitors N -Idimethylaminonaphthalene -5 -sulphonyl -Dtryptophan ethyl ester and the corresponding glycine derivative is much less good at pH 10 than at pH 8. The forces which are important for the binding of neutral inhibitors to ox-chymotrypsin are hydrophobic interactions (Knowles, 1965) and (largely for orientational purposes) hydrogen bonds (Ingles, Knowles & Tomlinson, 1966). Electrostatic interactions between enzyme and inhibitor cannot, of course, be relevant for the neutral inhibitor molecule.…”

Section: Resultsmentioning

confidence: 99%

The binding of inhibitors to α-chymotrypsin at alkaline pH

Johnson¹,

Knowles²

1967

Biochemical Journal

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1. The binding of the competitive inhibitor N-acetyl-d-tryptophan amide to alpha-chymotrypsin has now been studied at pH values up to 10.6, by the technique of equilibrium dialysis. 2. This binding depends on the ionization of a group on the free enzyme with apparent pK(a) 9.3 at 5 degrees . 3. This group is tentatively identified as that responsible for an enzyme conformation change at high pH values, on which the catalytic activity of the enzyme also depends.

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“…For example, k2 for TV-acetyl-L-tryptophanyl chymotrypsin is 2 X 104 times larger than k2 for TV-acetyl-D-tryptophanyl chymotrypsin. 33 The rate constant, k2, for the rate-determining step in the overall hydrolysis of either isomer of lid is greater than k0 at pH 7.0 and catalysis of the overall hydrolysis of lid by chymotrypsin does occur.…”

Section: Discussionmentioning

confidence: 98%