Identification of Glu-268 as the catalytic nucleophile of human lysosomal beta-galactosidase precursor by mass spectrometry.

نویسندگان

  • J D McCarter
  • D L Burgoyne
  • S Miao
  • S Zhang
  • J W Callahan
  • S G Withers
چکیده

Human lysosomal beta-galactosidase catalyzes the hydrolysis of beta-galactosides via a double displacement mechanism involving a covalent glycosyl enzyme intermediate. By use of the slow substrate 2,4-dinitrophenyl-2-deoxy-2-fluoro-beta-D-galactopyranoside, a glycosyl enzyme intermediate has been trapped on the enzyme. This has allowed the catalytic nucleophile to be identified as Glu-268 by peptic and tryptic digestion of the inactivated enzyme followed by high performance liquid chromatography-electrospray ionization tandem mass spectrometry of the peptide mixture. This glutamic acid is fully conserved in a sequence-related family of enzymes (Family 35), consistent with its essential role.

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عنوان ژورنال:
  • The Journal of biological chemistry

دوره 272 1  شماره 

صفحات  -

تاریخ انتشار 1997