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Chemical Studies of Structural Features in Staphylococcal Nuclease-T
Journal article   Open access   Peer reviewed

Chemical Studies of Structural Features in Staphylococcal Nuclease-T

Irwin M. Chaiken
The Journal of biological chemistry, v 247(7), pp 1999-2007
10 Apr 1972
DOI: https://doi.org/10.1016/S0021-9258(19)45482-3
PMID: 4335858
url
https://doi.org/10.1016/s0021-9258(19)45482-3View
Published, Version of Record (VoR) Open
url
https://doi.org/10.1016/S0021-9258(19)45482-3View
Published, Version of Record (VoR) Open

Abstract

Synthetically and chemically modified forms of staphylococcal nuclease-T′, the noncovalent complex of the polypeptide fragments nuclease-T-(6–48) (containing residues 6 through 48 of staphylococcal nuclease) and nuclease-T-(49–149) (containing residues 49 through 149 of nuclease), were prepared and characterized. Analogue forms of synthetic-(6–47) (the synthetic fragment corresponding to nuclease residues 6 through 47) were prepared by the solid phase method with variations from the normal sequence at residues lysine 9, glutamic acid 10, proline 11, alanine 12, threonine 13, leucine 14, isoleucine 15, isoleucine 18, and tyrosine 27. In addition, [3-nitro-l-tyrosyl 27]-nuclease-T-(6–48) and [3-amino-l-tyrosyl 27]-nuclease-T-(6–48) were obtained, respectively, by tetranitromethane treatment of nuclease-T-(6–48) and by subsequent reduction of the nitrated product. All of the above fragment derivatives were studied for the ability to associate with nuclease-T-(49–149) to generate enzymic activity. The results have allowed a description of the functions of the residues studied in nuclease-T′. This information has been interpreted further based on the present knowledge of the staphylococcal nuclease crystal structure (Arnone, A., Bier, C. J., Cotton, F. A., Hazen, E. E., Jr., Richardson, D. C., Richardson, J. S., AND Yonath, A. (1971) J. Biol. Chem. 246, 2302) and therein extended to predictions for intact nuclease as well. The studies reported indicate that the NH2-terminal residues 9 through 13 participate in several critical main chain and side chain hydrogen bonds, with loss of residues 6 through 13 causing complete destabilization of the nuclease-T′ complex. Tyrosine 27 helps to stabilize nuclease-T′ structure, probably by hydrophobic interactions as well as by side chain hydrogen bonding, whereas the nonpolar side chains of isoleucine 15 and isoleucine 18 are important for nuclease-T′ fragment association and stabilization of the active site region of the complex, respectively. The results are consistent with the view that nuclease and nuclease-T′ have similar detailed structural features in the regions studied.

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