Endoplasmic Reticulum Glucosidase II Is Composed of a Catalytic Subunit, Conserved from Yeast to Mammals, and a Tightly Bound Noncatalytic HDEL-containing Subunit*
- From the Department of Cell Biology, Yale University School of Medicine, P.O. Box 208002, New Haven, Connecticut 06520-8002
- ¶To whom all correspondence should be addressed. Tel.: 203-785-4315; Fax: 203-785-7226; E-mail: ari_helenius{at}qm.yale.edu.
Abstract
Trimming of glucoses from N-linked core glycans on newly synthesized glycoproteins occurs sequentially through the action of glucosidases I and II in the endoplasmic reticulum (ER). We isolated enzymatically active glucosidase II from rat liver and found that, in contrast with previous reports, it contains two subunits (α and β). Sequence analysis of peptides derived from them allowed us to identify their corresponding human cDNA sequences. The sequence of the α subunit predicted a soluble protein (104 kDa) devoid of known signals for residence in the ER. It showed homology with several other glucosidases but not with glucosidase I. Among the homologues, we identified a Saccharomyces cerevisiae gene, which we showed by gene disruption experiments to be the functional catalytic subunit of glucosidase II. The disrupted yeast strains had no detectable growth defect. The sequence of the β subunit (58 kDa) showed no sequence homology with other known proteins. It encoded a soluble protein rich in glutamic and aspartic acid with a putative ER retention signal (HDEL) at the C terminus. This suggested that the β subunit is responsible for the ER localization of the enzyme.
Footnotes
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↵‡ Fellow of The Jane Coffin Childs Memorial Fund for Medical Research.
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↵§ Supported by a postdoctoral fellowship from the Human Frontier Science Program Organization (LT-140/93).
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↵* This investigation has been aided by a grant from the The Jane Coffin Childs Memorial Fund for Medical Research and National Institutes of Health Grant GM 55972. The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵1 The abbreviations used are:
- ER
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endoplasmic reticulum
- DTT
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dithiothreitol
- PDI
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protein disulfide isomerase
- CPY
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carboxypeptidase Y
- PAGE
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polyacrylamide gel electrophoresis
- Endo H
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endo-β-N-acetylglucosaminidase H.
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- Received June 17, 1996.
- Revision received August 20, 1996.
- © 1996 by The American Society for Biochemistry and Molecular Biology, Inc.
