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. 1979 Apr;76(4):1566–1569. doi: 10.1073/pnas.76.4.1566

Detection and accumulation of tetrahedral intermediates in elastase catalysis.

A L Fink, P Meehan
PMCID: PMC383430  PMID: 36609

Abstract

Tetrahedral intermediates in the reaction of elastase with specific di- and tripeptide p-nitroanilide substrates have been detected, accumulated, and stabilized at high pH by using subzero temperatures and fluid aqueous/organic cryosolvents. The tetrahedral adducts are characterized by spectra with lambda max of 359 +/- 2 nm, compared with thata of 380 nm for p-nitroaniline and 315-320 nm for the substrates. The maximal concentration of intermediate that could be accumulated varied with the different substrates from 40 to 100% of the active enzyme present. The pH dependence of the reactions indicated that formation of the tetrahedral intermediates was rate-limiting at low pH (pK* = 7.0 at -39 degrees C) and that collapse to the acylenzymes was rate-determining at high pH. When corrected for the effect of temperature and cosolvent, the rate of intermediate formation was in good agreement with that measured at 25 degrees C in aqueous solution by stopped-flow techniques.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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