Role of cysteine residues in human plasma phospholipid transfer protein

Shi Jing Qu, Hui Zhen Fan, Cumhur Kilinc, Henry J. Pownall

Research output: Contribution to journalArticlepeer-review

7 Scopus citations

Abstract

Phospholipid transfer protein (PLTP) belongs to a family of human plasma lipid transfer proteins that bind to small amphophilic molecules. PLTP contains cysteines at residues 5, 129, 168, and 318. Bactericidal/permeability-increasing protein, which is a member of the same gene family, contains an essential disulfide bond between Cys135 and Cys175; these residues, which correspond to Cys!29 and Cys168 in PLTP, are conserved among all known members of the gene family. To identify the importance of these and the remaining cysteine residues to PLTP secretion and activity, each was replaced by a glycine by site-directed mutagenesis. The mutant as well as wild-type PLTP cDNAs were cloned into the mammalian expression vector pSV-SPORTl, and the PLTP cDNAs were transfected to COS-6 cells for expression. PLTP Cys129→Gly and PLTP CyS168→ Gly were secretion incompetent. Neither PLTP mass nor activity was detectable in cell lysates and culture medium. Relative to wild-type PLTP, PLTP Cyss → Gly and PLTP Cys318 → Gly exhibited similar specific activities but partially impaired PLTP synthesis and secretion. Intracellular PLTP appeared as two bands of 75 and 51 kDa corresponding to reported molecular masses for the glycosylated and nonglycosylated forms. The specific activities of PLTP Cys5→ Gly and PLTP Cys318 → Gly were similar in the cell lysates and medium, suggesting that glycosylation does not affect transfer activity.

Original languageEnglish (US)
Pages (from-to)193-198
Number of pages6
JournalJournal of Protein Chemistry
Volume18
Issue number2
DOIs
StatePublished - 1999

Keywords

  • Disulfide bonds
  • Lipid transfer
  • Lipoprotein metabolism
  • Site-directed mutagenesis

ASJC Scopus subject areas

  • Biochemistry
  • Analytical Chemistry
  • Organic Chemistry
  • Bioengineering

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