The protein synthesis factor EF-1Alpha of Mucor racemosus hyphae contains 8 or 9 methylated lysines per molecule, while the factor from assexual sporangiospores is non-methylated. During the course of spore germination the specific acitivity increases 6 to 9-fold. This increase in specific activity is accompanied by a constant level of mRNA and a level of EF-1Alpha protein that changes little during the germination period. By isolating and sequencing the EF-1Alpha gene, we found tht it is present in 3 copies in the Mucor genome, each in a different genomic context. This proposal is concerned with two aspects of EF-1Alpha: the role of methylation in """"""""activating"""""""" the factor, and the genetic organization, including the control of expression of each gene. We will study the effect of methylation on various EF-1Alpha activities in vitro, locate each methylated site within the protein and purify and study the EF-1Alpha methylases. We will further prepare gene-specific hybridization probes to assess the expression of each gene during several morphogenetic changes (e.g., spore germination, yeast-to-hypha conversions, and sporulation). We also propose to study the 5'flanking regions of each gene to assess the mechanisms controlling transcription.
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