Mono ADP-ribosylation reactions of proteins and guanidine derivatives can be carried out by specific bacterial toxins, e.g., cholera toxin and E. coli enterotoxin or by enzymes present in normal tissues. The exact function of these latter enzymes is not known and we will concentrate our efforts to establish the role of the enzyme we found in skeletal muscle. A comparison will be made with reactions catalyzed by cholera toxin to learn how these processes occur and what effects these reactions could have on other enzyme-catalyzed covalent modification reactions. Specifically, we plan to study the following topics: I. NAD-dependent Mono ADP-ribosyltransferase from Skeletal Muscle A. Enzyme Isolation B. Determination of Physical, Chemical, and Catalytic Properties of the Purified Enzyme C. Identification of Natural Protein Substrates II. The Process of Mono ADP-ribosylation A. The Kinetic Mechanism B. Specificity of the Reaction 1. Action on synthetic peptides 2. Action on guanyl hydrazones 3. Modification sites in proteins C. Active Site Directed Reagents D. Reversibility of the Process III. Consequences of ADP-ribosylation Reactions A. Effects on Other Enzyme Catalyzed Covalent Modification Reactions B. On Reactions Occurring in the Glycogen Particle, Sarcoplasmic Reticulum and Sarcolemma
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