The growth-related 28-kilodalton protein in germinating wheat. Use of peptide mapping to identify cryptic forms in cell-free extracts and protein-synthesizing systems

Abstract

Characteristic peptide maps have been prepared and used to identify cryptic forms of the methionine-labeled 28-kilodalton (28-kd) protein (the G protein) which marks onset of growth in germinating wheat embryos. Peptide mapping of material eluted from areas detected as "bands" in autoradiograms of one-dimensional Laemmli-type gels and from areas detected as "spots" in autoradiograms of two-dimensional O'Farrell-type gels has led to identification of a counterpart to G among methionine-labeled products of cell-free synthesis. Identification of its cell-free counterpart has made it possible to show that translatable mRNA for the G protein, like G itself, only begins to appear between 5 and 10 h after onset of water uptake by dry wheat embryos. Peptide mapping of methionine- or proline-labeled material eluted from an area detected as a conspicuous "band" in autoradiograms of one-dimensional Laemmli-type gels has uncovered the existence of an unusual (apparently complexed) form of G in extracts of germinating but not dry wheat embryos. Unique among the hundreds of prominent proteins made by germinating wheat embryos, G exists as part of a thermolabile "150-kd complex" which is refractory to dismutation in a reducing environment containing sodium dodecyl sulphate (SDS). Presence or absence of the G complex constitutes the only conspicuous difference between SDS–polyacrylamide distributions of newly synthesized proteins derived from unheated as opposed to heated extracts of germinating embryos. Analysis of extracts prepared at different times after onset of water imbibition by dry wheat embryos shows that time course of emergence of the G complex, detected in unheated extracts, and of the G protein, detected in heated extracts, is the same. By fractional electroelution and silver staining of protein, it has been possible to show that G may be the principal if not the only protein component of the G complex.