A Caltech Library Service

Sulfhydryl Modification of the Yeast Wbp1p Inhibits Oligosaccharyl Transferase Activity

Pathak, Rahul and Hendrickson, Tamara L. and Imperiali, Barbara (1995) Sulfhydryl Modification of the Yeast Wbp1p Inhibits Oligosaccharyl Transferase Activity. Biochemistry, 34 (13). pp. 4179-4185. ISSN 0006-2960. doi:10.1021/bi00013a005.

Full text is not posted in this repository. Consult Related URLs below.

Use this Persistent URL to link to this item:


Chemical labeling of the multimeric Saccharomyces cerevisiae oligosaccharyl transferase indicates that the 48 kDa Wbplp subunit is an integral component of the catalytically active enzyme. The enzyme was purified following chromatography on concanavalin A agarose, heparin agarose, Q-Sepharose, and hydroxyapatite media. The enzyme activity copurified with a tetrameric complex of polypeptide subunits. Two of the subunits have been identified as the yeast proteins Wbplp and Swplp by amino-teminal residue sequencing. A third subunit was identified as a variably glycosylated polypeptide near 64 kDa; preliminary amino acid sequencing showed no identity to known yeast proteins. Modification of a cysteine residue by the reagent methyl methanethiolsulfonate (MMTS) caused time-dependent and concentration-dependent inactivation of the enzyme. To identify the modified subunit of the transferase complex, the labeling reagent S-[(N-biotinoy1amino)ethyll methanethiolsulfonate (BMTS) was synthesized. Like MMTS, BMTS inactivated the oligosaccharyl transferase in a time-dependent manner. Additionally, incubation with the substrate (dolichylpyrophosphory1)-N,N'-diacetylchitobiose [Dol-PP(GlcNAc)_2] protected the enzyme from BMTS inactivation. When the purified enzyme complex was incubated with BMTS, Wbp lp alone was specifically labeled, thereby associating this subunit with catalysis and the binding of the dolichylpyrophosphoryl oligosaccharide substrate in the transferase reaction.

Item Type:Article
Related URLs:
URLURL TypeDescription
Additional Information:© 1995 American Chemical Society. Published in print 4 April 1995. This work was supported by the National Institutes of Health Grant GM39334, the Alfred P. Sloan Foundation, the Camille and Henry Dreyfus Teacher Scholar Program, and Zeneca, Inc. R.P. was supported by a National Science Foundation predoctoral fellowship, and T.L.H. is supported by the NIH Predoctoral Biotechnology Training Grant GM08346. We thank Phillips W. Robbins and Janet W. Zimmennan for providing the yeast strain used for the purification and Markus Aebi for providing antiserum to the Wbplp protein. Contribution number 9001
Funding AgencyGrant Number
Alfred P. Sloan FoundationUNSPECIFIED
Camille and Henry Dreyfus FoundationUNSPECIFIED
NSF Predoctoral FellowshipUNSPECIFIED
NIH Predoctoral FellowshipGM08346
Other Numbering System:
Other Numbering System NameOther Numbering System ID
Caltech Division of Chemistry and Chemical Engineering9001
Issue or Number:13
Record Number:CaltechAUTHORS:20180205-080953639
Persistent URL:
Official Citation:Sulfhydryl Modification of the Yeast Wbp1p Inhibits Oligosaccharyl Transferase Activity Rahul Pathak, Tamara L. Hendrickson, and Barbara Imperiali Biochemistry 1995 34 (13), 4179-4185 DOI: 10.1021/bi00013a005
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:84664
Deposited By: Ruth Sustaita
Deposited On:05 Feb 2018 23:43
Last Modified:15 Nov 2021 20:21

Repository Staff Only: item control page