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PMID: 10357807 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

Functional glycan-free adhesion domain of human cell surface receptor CD58: design, production and NMR studies.

The EMBO journal ·Vol. 18 ·No. 11 ·1999-06-01 ·Pages 2941-9

Sun ZY, Dötsch V, Kim M, Li J, Reinherz EL, Wagner G

Abstract

A general strategy is presented here for producing glycan-free forms of glycoproteins without loss of function by employing apolar-to-polar mutations of surface residues in functionally irrelevant epitopes. The success of this structure-based approach was demonstrated through the expression in Escherichia coli of a soluble 11 kDa adhesion domain extracted from the heavily glycosylated 55 kDa human CD58 ectodomain. The solution structure was subsequently determined and binding to its counter-receptor CD2 studied by NMR. This mutant adhesion domain is functional as determined by several experimental methods, and the size of its binding site has been probed by chemical shift perturbations in NMR titration experiments. The new structural information supports a 'hand-shake' model of CD2-CD58 interaction involving the GFCC'C" faces of both CD2 and CD58 adhesion domains. The region responsible for binding specificity is most likely localized on the C, C' and C" strands and the C-C' and C'-C" loops on CD58.

MeSH Terms
Amino Acid Sequence Binding Sites CD2 Antigens/chemistry,metabolism CD58 Antigens/biosynthesis,chemistry,genetics,metabolism Escherichia coli/genetics Glycosylation Humans Models, Molecular Molecular Sequence Data Molecular Weight Mutation Nuclear Magnetic Resonance, Biomolecular Peptide Fragments/biosynthesis,chemistry,genetics,metabolism Polysaccharides/chemistry,genetics Protein Conformation Protein Engineering Protein Structure, Secondary Recombinant Proteins/biosynthesis,chemistry,metabolism Solubility
Chemicals
CD2 Antigens CD58 Antigens Peptide Fragments Polysaccharides Recombinant Proteins
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Sun Z Y
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA, USA.
Dötsch V
Kim M
Li J
Reinherz E L
Wagner G
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Article Info
Journal
The EMBO journal
Abbr.
EMBO J
ISSN
0261-4189
DOI
10.1093/emboj/18.11.2941
Published
1999-06-01
Pages
2941-9
Language
English
Region
England
NLM ID
8208664
PMCID
PMC1171376
Subset
IM
Grants
NIAID NIH HHS · AI21226 · United States
NIAID NIH HHS · AI37581 · United States
Databases
PDB
1CI5
External Links
PubMed source DOI: 10.1093/emboj/18.11.2941 PMC Full Text View journal details
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