Metal Ion Dependence of Oligosaccharyl Transferase: Implications for Catalysis
- Creators
- Hendrickson, Tamara L.
- Imperiali, Barbara
Abstract
Oligosaccharyl transferase activity exhibits an absolute requirement for certain divalent metal cations. Studies with reconstituted enzyme suggest a preference for metal ions that can adopt an octahedral coordination geometry. In order to gain insight into the specific role of the metal cation in catalysis, we have investigated the influence of the metal cofactor on catalytic tumover of the tripeptide substrate Bz- Asn-Leu-Thr-NHMe (1) and a closely related sulfur-containing analog, Bz-Asn(yS)-Leu-Thr-NHMe (2). The metal ion substitution studies reveal that 1 is effectively tumed over in the presence of several metal ions (Mn^(2+), Fe^(2+), Mg^(2+), and Ca^(2+)). In contrast, 2 is only glycosylated in the presence of the thiophilic metal cations manganese and iron. When the enzyme is reconstituted with the oxophilic cations magnesium and calcium, 2 shows minimal substrate behavior. With the amide substrate 1, the distinct preference for manganese over magnesium may argue against direct coordination of the metal to the lipid-linked substrate pyrophosphate moiety. This fact, together with the comparative studies with asparagine- and thioasparagine-containing tripeptides, implicates the metal cofactor in a role that places it proximal to the peptide binding site.
Additional Information
© 1995 American Chemical Society. Published in print 25 July 1995. This work was supported by National Institutes of Health Grant GM39334, the Alfred P. Sloan Foundation, the Camille and Henry Dreyfus Teacher Scholar Program, and Zeneca, Inc. T.L.H. is supported by NIH Predoctoral Biotechnology Training Grant GM08346. Contribution no. 9046.Attached Files
Supplemental Material - bi00029a020_si_001.pdf
Supplemental Material - bi9444.pdf
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Additional details
- Eprint ID
- 84662
- DOI
- 10.1021/bi00029a020
- Resolver ID
- CaltechAUTHORS:20180205-073724083
- GM39334
- NIH
- Alfred P. Sloan Foundation
- Camille and Henry Dreyfus Foundation
- Zeneca
- GM08346
- NIH Predoctoral Fellowship
- Created
-
2018-02-05Created from EPrint's datestamp field
- Updated
-
2021-11-15Created from EPrint's last_modified field
- Other Numbering System Name
- Caltech Division of Chemistry and Chemical Engineering
- Other Numbering System Identifier
- 9046