Send to

Choose Destination
FEBS Lett. 2007 May 29;581(13):2441-6. Epub 2007 Apr 30.

Identification of the catalytic nucleophile in Family 42 beta-galactosidases by intermediate trapping and peptide mapping: YesZ from Bacillus subtilis.

Author information

Department of Chemistry, University of British Columbia, 2036 Main Mall, Vancouver, BC, Canada.


The mechanism-based inhibitor 2,4-dinitrophenyl 2-deoxy-2-fluoro-beta-d-galactopyranoside (DNP2FGal) was used to inactivate the Family 42 beta-galactosidase (YesZ) from Bacillus subtilis via the trapping of a glycosyl-enzyme intermediate, thereby tagging the catalytic nucleophile in the active site. Proteolytic digestion of the inactivated enzyme and of a control sample of unlabeled enzyme, followed by comparative high-performance liquid chromatography and mass spectrometric analysis identified a labelled peptide of the sequence ETSPSYAASL. These data, combined with sequence alignments of this region with representative members of Family 42, unequivocally identify the catalytic nucleophile in this enzyme as Glu-295, thereby providing the first direct experimental proof of the identity of this residue within Family 42.

[Indexed for MEDLINE]
Free full text

Supplemental Content

Full text links

Icon for Wiley
Loading ...
Support Center