Studies on adenosine triphosphate transphosphorylases. XVIII. Synthesis and preparation of peptides and peptide fragments of rabbit muscle ATP-AMP transphosphorylase (adenylate kinase) and their nucleotide-binding properties |
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| Authors: | Stephen A Kuby Minoru Hamada Mark S Johnson Geoffrey A Russell Michael Manship Richard H Palmieri Gerald Fleming David S Bredt and Albert S Mildvan |
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| Institution: | (1) Laboratory for the Study of Hereditary and Metabolic Disorders and Departments of Biochemistry and Medicine, University of Utah, 84108 Salt Lake City, Utah;(2) Present address: Department of Hygiene, Miyazaki Medical College, Miyazaki, Japan;(3) Present address: Department of Microbiology, Loma Linda University, Loma Linda, California;(4) Present address: Division of Analytical Sciences, Kodak Research Laboratories, Rochester, New York;(5) Present address: Beckman Instruments, Inc., 84308 Palo Alto, California;(6) Department of Biological Chemistry, Johns Hopkins School of Medicine, 21205 Baltimore, Maryland |
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| Abstract: | Two peptide fragments, derived from the head and tail of rabbit muscle myokinase, were found to possess remarkable and specific ligand-binding properties (Hamadaet al., 1979).By initiating systematic syntheses and measurements of equilibrium substrate-binding properties of these two sets of peptides, or portions thereof, which encompass the binding sites for (a) the magnesium complexes of the nucleotide substrates (MgATP2– and MgADP–) and (b) the uncomplexed nucleotide substrates (ADP3– and AMP2–) of rabbit muscle myokinase, some of the requirements for binding of the substrates to ATP-AMP transphosphorylase are being deduced and chemically outlined. One requirement for tight nucleotide binding appears to be a minimum peptide length of 15–25 residues. In addition, Lys-172 and/or Lys-194 may be involved in the binding of  AMP.The syntheses are described as a set of peptides corresponding to residues 31–45, 20–45, 5–45, and 1–45, and a set of peptides corresponding to residues 178–192, 178–194, and 172–194 of rabbit muscle adenylate kinase. The ligand-binding properties of the first set of synthetic peptides to the fluorescent ligands: MgATP/ATP and MgADP/ADP are quantitatively presented in terms of their intrinsic dissociation constants (K d) and values ofN (maximal number of moles bound per mole of peptide); and compared with the peptide fragment MT-I (1–44) obtained from rabbit muscle myokinase (Kubyet al., 1984) and with the native enzyme (Hamadaet al., 1979). In addition, the values ofN andKd are given for the second set of synthetic peptides to the fluorescent ligands AMP and ADP as well as for the peptide fragments MT-XII(172–194) and CB-VI(126–194) (Kuby et al., 1984) and, in turn, compared with the native enzyme.A few miscellaneous dissociation constants which had been derived kinetically are also given for comparison (e.g., theK
i for AMP and the value of
obtained for the native enzyme) (Hamada and Kuby, 1978), and theK'd measured for Cr3+ and the synthetic peptide I1–45 (Fryet al., 1985b).Paper XVII of this series is Kubyet al. (1983). |
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| Keywords: | ATP-AMP transphosphorylase adenylate kinase myokinase nucleotide-binding peptides and peptide fragments ligand binding peptide synthesis |
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