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Biochemistry (Moscow)

, Volume 74, Issue 2, pp 194–200 | Cite as

Dependence of conformation of D3/D4 domains of human CD4 on glycosylation and membrane attachment

  • L. F. LidemanEmail author
  • R. A. Gibadulin
Article
  • 60 Downloads

Abstract

Conformational dynamics of human T-helper cell receptor protein CD4 has been studied with the help of monoclonal antibody (mAb) T6. The mAb T6 discriminates between s- and m-forms of CD4 and recognizes a specific conformation of the soluble (s) form of CD4 including the first nine amino acids of CD4 transmembrane sequence. However, change of tryptophan for serine in position 2 in this sequence destabilizes the T6-type conformation. By enzymatic deglycosylation and deletions of glycosylation sites, we show that T6-type conformation depends on glycosylation in both sites (Asn271 and Asn300). We show also that the sugars are not involved in direct binding to the antibody but stabilize the D3/D4 local conformation. Deglycosylated forms of sCD4 in vivo acquire a specific conformation similar to the wild type sCD4, which however cannot be restored after denaturation/renaturation under conditions of non-reducing Western blot. This observation indicates that the correct protein folding needs chaperone assistance and cannot be achieved in vitro. Completely non-glycosylated sCD4 is synthesized and secreted into the growth medium. In the medium, this mutant appears to be unstable and aggregates during time. In a contrast to soluble CD4, mutations in glycosylation sites abrogate expression of membrane CD4, thus demonstrating a different secretion pathways for soluble and membrane proteins.

Key words

CD4 autoimmune epitope glycosylation protein folding HIV-1 

Abbreviations

a.a.

amino acid residue

IL

interleukin

mAb

monoclonal antibody

MHC

major histocompatibility complex

PBS

phosphate buffered saline

sCD4 and mCD4

soluble and membrane forms of CD4, respectively

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Copyright information

© Pleiades Publishing, Ltd. 2009

Authors and Affiliations

  1. 1.Ivanovsky Institute of VirologyRussian Academy of Medical SciencesMoscowRussia

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