Proteomic analysis of brush-border membrane vesicles isolated from purified proximal convoluted tubules

Scott J. Walmsley, Corey Broeckling, Ann Hess, Jessica Prenni, Norman P. Curthoys

Research output: Contribution to journalArticlepeer-review

20 Scopus citations

Abstract

The renal proximal convoluted tubule is the primary site of water, electrolyte and nutrient reabsorption and of active secretion of selected molecules. Proteins in the apical brush-border membrane facilitate these functions and initiate some of the cellular responses to altered renal physiology. The current study uses two-dimensional liquid chromatography/mass spectrometry to compare brush border membrane vesicles isolated from rat renal cortex (BBMVCTX) and from purified proximal convoluted tubules (BBMVPCT). Both proteomic data and Western blot analysis indicate that the BBMVCTX contain apical membrane proteins from cortical cells other than the proximal tubule. This heterogeneity was greatly reduced in the BBMVPCT. Proteomic analysis identified 193 proteins common to both samples, 21 proteins unique to BBMVCTX, and 57 proteins unique to BBMVPCT. Spectral counts were used to quantify relative differences in protein abundance. This analysis identified 42 and 50 proteins that are significantly enriched (p values ≤0.001) in the BBMVCTX and BBMVPCT, respectively. These data were validated by measurement of γ-glutamyltranspeptidase activity and by Western blot analysis. The combined results establish that BBMVPCT are primarily derived from the proximal convoluted tubule (S1 and S2 segments), whereas BBMVCTX include proteins from the proximal straight tubule (S3 segment). Analysis of functional annotations indicated that BBMVPCT are enriched in mitochondrial proteins and enzymes involved in glucose and organic acid metabolism. Thus the current study reports a detailed proteomic analysis of the brush-border membrane of the rat renal proximal convoluted tubule and provides a database for future hypothesis-driven research.

Original languageEnglish (US)
Pages (from-to)F1323-F1331
JournalAmerican Journal of Physiology - Renal Physiology
Volume298
Issue number6
DOIs
StatePublished - Jun 2010
Externally publishedYes

Keywords

  • Apical membrane
  • Functional annotations
  • Membrane proteins
  • Rat kidney
  • Spectral counting

Fingerprint Dive into the research topics of 'Proteomic analysis of brush-border membrane vesicles isolated from purified proximal convoluted tubules'. Together they form a unique fingerprint.

Cite this