Chemical analysis of processing of spiralin, the major lipoprotein of Spiroplasma melliferum

Arch Microbiol. May-Jun 2000;173(5-6):339-45. doi: 10.1007/s002030000145.

Abstract

The plasma membrane of Spiroplasma melliferum contains a major membrane-associated lipoprotein called spiralin. In this study, the processing pathway of spiralin was investigated by chemical analysis of the purified protein and by using [35S]cysteine, [35S]methionine, [14C]myristic acid (14C-14:0), [14C]palmitic acid (14C-16:0), and globomycin. SDS-PAGE analysis of membrane proteins showed the leader peptide cleavage of prospiralin and provided evidence for an apparent selectivity in the acylation: the unprocessed protein was labelled with 14C-16:0 only (O-ester-linked acyl chains), and the mature form with both 14C-labelled fatty acids (O-ester-linked + amide-linked chains). Chemical analysis of the purified protein revealed that spiralin contains S-glycerylcysteine and is covalently modified with two O-ester-linked acyl chains and one amide-linked fatty acid chain. However, a specific selectivity in the O- and the N-acylations was not confirmed; palmitate and stearate were the major components. The amounts of O-ester- and amide-linked acyl chains, the resistance to Edman degradation and the presence of S-glycerylcysteine together indicate that spiralin is a "classical" lipoprotein (i.e. is triacylated) and is probably processed by a mechanism similar to that described for gram-negative eubacteria. On the basis of these findings, a biogenesis pathway for spiralin is proposed.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acids / analysis
  • Bacterial Outer Membrane Proteins / chemistry*
  • Bacterial Outer Membrane Proteins / metabolism*
  • Culture Media
  • Cysteine / analogs & derivatives
  • Cysteine / analysis
  • Electrophoresis, Polyacrylamide Gel
  • Fatty Acids / analysis
  • Membrane Proteins / analysis
  • Protein Processing, Post-Translational*
  • Sequence Analysis, Protein
  • Spiroplasma / chemistry
  • Spiroplasma / growth & development
  • Spiroplasma / metabolism*
  • Sulfur Radioisotopes / metabolism

Substances

  • Amino Acids
  • Bacterial Outer Membrane Proteins
  • Culture Media
  • Fatty Acids
  • Membrane Proteins
  • Sulfur Radioisotopes
  • spiralin
  • glycerylcysteine
  • Cysteine