The long range goals of the proposed research is to understand how specificity signals encoded into protein sequences direct the co- and posttranstational processing of the linear polyamino acid products specified by the genetic code. It is becoming increasingly clear that co-/post-translational protein modification reactions represent key steps in the biosynthesis of biologically active proteins and in their transport to the appropriate compartments of action; our understanding of these reactions will be an important component in elucidating the genetic lesions in many inherited diseases and an absolute requirement for the full utilization of recombinant technology whenever proteins requiring extensive processing are produced in surrogate hosts lacking all or part of the appropriate processing apparatus. Two co-/post- translational reactions will be studied. The first, Glycan processing and recognition in glycoproteins, explores the fundamental question of how the protein matrix on which the common N-liked precursor glycan is located regulates the processing steps leading to different glycan products, and also how it affects the display of the glycan units in the interaction with receptors. Using neoglycoprotein models, the problem will be tackled by evaluating the substrate and ligand properties of well-defined free glycans in direct comparison with the same glycans associated with proteins in different types of complexes and with different glycan-protein stoichiometry. The second, the N-terminal processing of eukaryotic proteins, focuses on the reactions and the specificity signals involved in the well- established removal of the initiator methionine and the acylation of the new N-terminals amino acid in a large number of eukaryotic proteins. Building on the discovery of a muscle enzyme which catalyzes the specific removal of N- acetylmethionine from the N-terminus of short and long peptides, the proposed studies will focus on the enzymology of N-terminal trimming and acetylation in muscle and in yeast. The enzymes will be purified by conventional methods, and their specificity assessed by the use of different peptide (protein) substrates, the selection of which will be based on known acetylated and nonacetylated sequences in the data banks.
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