Display Settings:

Format

Send to:

Choose Destination

    Nature. 2001 Aug 23;412(6849):835-8.

    Catalysis by hen egg-white lysozyme proceeds via a covalent intermediate.

    Vocadlo DJ, Davies GJ, Laine R, Withers SG.

    Protein Engineering Network of Centres of Excellence and the Department of Chemistry, University of British Columbia, Vancouver, British Columbia V6T 1Z1, Canada.

    Hen egg-white lysozyme (HEWL) was the first enzyme to have its three-dimensional structure determined by X-ray diffraction techniques. A catalytic mechanism, featuring a long-lived oxocarbenium-ion intermediate, was proposed on the basis of model-building studies. The 'Phillips' mechanism is widely held as the paradigm for the catalytic mechanism of beta-glycosidases that cleave glycosidic linkages with net retention of configuration of the anomeric centre. Studies with other retaining beta-glycosidases, however, provide strong evidence pointing to a common mechanism for these enzymes that involves a covalent glycosyl-enzyme intermediate, as previously postulated. Here we show, in three different cases using electrospray ionization mass spectrometry, a catalytically competent covalent glycosyl-enzyme intermediate during the catalytic cycle of HEWL. We also show the three-dimensional structure of this intermediate as determined by X-ray diffraction. We formulate a general catalytic mechanism for all retaining beta-glycosidases that includes substrate distortion, formation of a covalent intermediate, and the electrophilic migration of C1 along the reaction coordinate.

    PMID: 11518970 [PubMed - indexed for MEDLINE]

    Supplemental Content

    Click here to read

    Structures reported by this article