Role of zinc in the structure and function of ssDNA binding proteins
Gene 32 protein (g32P), the ssDNA binding protein required for replication, recombination and translational control in the phage T4 life cycle contains 1 g at Zn per mol bound in a tetrahedral complex to -S/sup -/ ligands. Chemical modification and spectroscopic evidence suggest binding to Cys 77, His 81, Cys 87 and Cys 90. The Zn-binding domain is at the N-terminal end of AA residues 72 to 116 which contains 6 of the 8 Tyr residues in g32P, shown by /sup 1/H-NMR to be involved in deoxynucleotide binding. Limited proteolysis of g32P with trypsin removes residues 1-21 and 254-301 leaving a trypsin-resistant core, g32P. The latter retains high affinity for a single site nucleotide lattice, but has lost cooperative binding to DNA. Removal of Zn from native g32P renders the molecule susceptible to proteolysis and the core is hydrolyzed to small peptides. Rebinding of Zn restores the core stability. Apo g32P binds 3 orders of magnitude less tightly to ssDNA and cannot melt polyd(AT) at 150 mM NaCl. Loss of binding affinity is primarily due to loss of cooperative protein-protein interactions accompanying Zn removal. Thus, both the N-terminal domain and the Zn-binding domain are required for cooperative binding to DNA. G5P from fd and ssB from E. coli do not contain Zn, but small basic proteins, products of the gag gene of retroviruses, e.g., p10 from MuLV, p15 from HTLVI and p15 from HTLV-III contain tandem repeats of a -C-X/sub 2/-C-X/sub 4/-H-X/sub 4/-C- sequence similar to the Zn-binding sequence found in g32P.
- Research Organization:
- Yale Univ., New Haven, CT
- OSTI ID:
- 6018035
- Report Number(s):
- CONF-870644-
- Journal Information:
- Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Vol. 46:6; Conference: 78. annual meeting of the American Society of Biological Chemists conference, Philadelphia, PA, USA, 7 Jun 1987
- Country of Publication:
- United States
- Language:
- English
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59 BASIC BIOLOGICAL SCIENCES
PROTEINS
BIOLOGICAL FUNCTIONS
MOLECULAR STRUCTURE
ZINC
BIOCHEMICAL REACTION KINETICS
BIOLOGICAL EFFECTS
AFFINITY
AMINO ACID SEQUENCE
DNA
DNA REPLICATION
GENE RECOMBINATION
LIGANDS
NMR SPECTRA
NUCLEAR MAGNETIC RESONANCE
ELEMENTS
FUNCTIONS
KINETICS
MAGNETIC RESONANCE
METALS
NUCLEIC ACID REPLICATION
NUCLEIC ACIDS
ORGANIC COMPOUNDS
REACTION KINETICS
RESONANCE
SPECTRA
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