Crystalline arrays of the Escherichia coli sn-glycerol-3-phosphate acyltransferase, an integral membrane protein.

Journal Article

The gene encoding the Escherichia coli sn-glycerol-3-phosphate acyltransferase, plsB, was inserted into hybrid plasmids under transcriptional control of the lambda PL and tac promoters. Enzymatic activities 35-50-fold above wild type and a large increase in glycerol-P acyltransferase polypeptide were obtained. Thin section electron microscopy of the cells overproducing the glycerol-P acyltransferase revealed 235-245-A diameter tubular structures associated with the cytoplasmic membrane. These structures were released from the cell by osmotic lysis and purified on Matrex Gel Green A. Subsequent sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated that the dominant protein constituent of the tubules was the glycerol-P acyltransferase. Analysis of tubule-enriched fractions isolated by differential centrifugation revealed a decreased phospholipid to protein ratio as compared to total and cytoplasmic membrane fractions. At high magnification, negative stained tubules displayed ordered arrays of stain-excluding components projecting 50-60 A from the cytoplasmic surface. Optical diffraction patterns from the micrographs contained intense layer lines at (1/78 A) and (1/39 A) along the tubule axis and a prominent spot at (1/62 A) near the equator. From compositional and structural data, 18-37% of the polypeptide volume is estimated to lie within the hydrophobic domain of the tubule membrane.

Full Text

Duke Authors

Cited Authors

  • Wilkison, WO; Walsh, JP; Corless, JM; Bell, RM

Published Date

  • July 25, 1986

Published In

Volume / Issue

  • 261 / 21

Start / End Page

  • 9951 - 9958

PubMed ID

  • 3525537

International Standard Serial Number (ISSN)

  • 0021-9258

Language

  • eng

Conference Location

  • United States