Analyses of genetic variants of l-glycerol-3-phosphate dehydrogenase in Drosophila melanogaster by two-dimensional gel electrophoresis and immunoelectrophoresis

Chi Yu Lee, David Niesel, Glenn C. Bewley

Research output: Contribution to journalArticle

9 Scopus citations

Abstract

A protein spot corresponding to l-glycerol-3-phosphate dehydrogenase (α-GPDH, E.C. 1.1.1.8, NAD+ oxidoreductase) has been identified on a two-dimensional gel (isoelectric focusing-SDS gel) containing up to 150 stained protein spots from a crude Drosophila homogenate. Preliminary identification of the α-GPDH spot was made by including a suitable amount of purified Drosophila α-GPDH in crude fly homogenates prior to electrophoresis and observing an intensity enhancement of the corresponding protein spot on the gels. When three purified electrophoretic variants (slow, fast, and ultrafast) were mixed and analyzed by two-dimensional gel electrophoresis, horizontal displacements of the three protein spots were observed. Immunoprecipitation of the enzyme prior to electrophoresis and gene mapping further confirmed the identity of the α-GPDH protein spot. The α-GPDH spot can also be detected by autoradiography of a two-dimensional gel from a single fly extract, where it has been estimated to constitute 0.5-1% of the total soluble protein. Mutants which express no apparent α-GPDH activity were analyzed by two-dimensional gels and immunoelectrophoresis in an attempt to identify and characterize the inactive proteins. It is suggested that these techniques provide a powerful tool for the analysis of CRM+-null activity mutants of a specific gene-enzyme system.

Original languageEnglish (US)
Pages (from-to)1003-1018
Number of pages16
JournalBiochemical Genetics
Volume18
Issue number9-10
DOIs
StatePublished - Oct 1 1980
Externally publishedYes

    Fingerprint

Keywords

  • immunoelectrophoresis
  • two-dimensional electrophoresis
  • α-GPD null mutants

ASJC Scopus subject areas

  • Ecology, Evolution, Behavior and Systematics
  • Biochemistry
  • Molecular Biology
  • Genetics

Cite this