Interaction of spermatid-specific protein TP2 with nucleic acids, in vitro. A comparative study with TP1.
TP2 was purified from rat testes employing a gentle method involving differential salt extraction of the sonication-resistant spermatid nuclei. The nucleic acid binding properties of TP2 were studied by fluorescence quenching, thermal denaturation, circular dichroism techniques and compared with those of TP1 (Singh, J., and Rao, M. R. S. (1987) J. Biol. Chem. 262, 734-740). The tyrosine fluorescence of TP2 was quenched upon binding to double-stranded and denatured DNA and poly(rA). The apparent association constants for binding of TP2 to these nucleic acids were calculated from the fluorescence quenching data, obtained at 50 mM NaCl, and found to be 1.63 x 10(5) M-1, 6.5 x 10(5) M-1, and 7.3 x 10(5) M-1, respectively. Thermal denaturation studies of calf thymus DNA and its complexes with TP2 showed that at 1 mM NaCl, TP2 shifted the Tm from 53 degrees C to 62-67 degrees C, while at 50 mM NaCl, the Tm was shifted from 72 to 78 degrees C suggesting that TP2 is a DNA stabilizing protein. Circular dichroism studies of TP1.DNA and TP2.DNA complexes have revealed that TP2 has a better DNA condensing property than TP1. Furthermore, in contrast to TP1, TP2 does not destabilize in vitro the compactness of liver nucleosome core particles. The DNA binding properties of TP1 and TP2 have been discussed in relation to the significance of their transient appearance during mammalian spermiogenesis.
Baskaran
R
R
Department of Biochemistry, Indian Institute of Science, Bangalore.
Rao
M R
MR
D000595:Amino Acid Sequence; D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D002850:Chromatography, Gel; D002868:Chromosomal Proteins, Non-Histone / Q000302:isolation & purification / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D006657:Histones / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000378:metabolism; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D009707:Nucleosomes / Q000378:metabolism; D051381:Rats; D012689:Sequence Homology, Nucleic Acid; D013050:Spectrometry, Fluorescence; D013087:Spermatids / Q000378:metabolism*; D013737:Testis / Q000378:metabolism
D002868:Chromosomal Proteins, Non-Histone; D006657:Histones; D009707:Nucleosomes; C053886:spermatid transition proteins; D004247:DNA