Bacterial and viral sialidases: Contribution of the conserved active site glutamate to catalysis

Jefferson Chan, Jacqueline N. Watson, April Lu, Viviana C. Cerda, Thor J. Borgford, Andrew J. Bennet

Research output: Contribution to journalArticlepeer-review


Mutagenesis of the conserved glutamic acid of influenza type A (E277) and Micromonospora viridifaciens (E260) sialidases was performed to probe the contribution of this strictly conserved residue to catalysis. Kinetic studies of the E260D and E260C M. viridifaciens mutant enzymes reveal that the overall mechanism of action has not changed. That is, the mutants are retaining sialidases in which glycosylation and deglycosylation are rate-limiting for k cat/K m and k cat, respectively. The solvent kinetic isotope effect and proton inventory on k cat for the E260C mutant sialidase provide strong evidence that the newly installed cysteine residue provides little catalytic acceleration. The results are consistent with the conserved aspartic acid residue (D92) becoming the key general acid/base residue in the catalytic cycle. In addition, the E277D mutant influenza type A sialidase is catalytically active toward 4-nitrophenyl α-D-sialoside, although no measurable hydrolysis of natural substrates was observed. Thus, mutating the glutamate residue (E277) to an aspartate increases the activation free energy of hydrolysis for natural substrates by >22 kJ/mol.

Original languageEnglish (US)
Pages (from-to)433-441
Number of pages9
Issue number1
StatePublished - Jan 10 2012
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry


Dive into the research topics of 'Bacterial and viral sialidases: Contribution of the conserved active site glutamate to catalysis'. Together they form a unique fingerprint.

Cite this