Characterization of poly C preferential ribonuclease from chicken liver.
Poly C preferential RNase previously reported by Levy and Karpetsky [J. Biol. Chem. 255, 2153-2159 (1980)] and Miura et al. [Chem. Pharm. Bull. 32, 4053-4060 (1984)] was extensively purified from chicken liver to homogeneity as determined by SDS-PAGE (RNase CL2). The poly C preference over poly U was slightly higher than that of bovine pancreatic RNase A. However, the kinetic constants for 8 dinucleoside phosphates, CpY and UpY (Y = one of A, G, U, and C) as substrates showed that RNase CL2 was preferential for cytidylic acid, but less so than RNase A, and the influence of Y base on the rate of hydrolysis of CpY or UpY was less marked than in the case of RNase A. The primary structure of RNase CL2 was determined. The molecular weight calculated from the sequence was 13,420. Comparison of the amino acid sequence of RNase CL2 with those of other vertebrate RNases showed that RNase CL2 is a member of the RNase A family, but is not a non-secretory RNase. It retains 3 disulfide bridges of RNase A, but Cys65-Cys72 of RNase A is missing. As for the active site, the amino acid residues of the P0 and P1 sites of RNase A are completely conserved. Among the B1 site components, Thr45 (RNase A numbering) is conserved, but Phe120 and Ser123 are substituted by Leu and Thr, respectively. Among the B2 site residues, Gln69, Asn71, and Glu111 are substituted by other amino acids.
Hayano
K
K
Department of Microbiology, Hoshi College of Pharmacy, Tokyo.
Iwama
M
M
Sakamoto
H
H
Watanabe
H
H
Sanda
A
A
Ohgi
K
K
Irie
M
M
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002645:Chickens; D002851:Chromatography, High Pressure Liquid; D017124:Conserved Sequence; D003568:Cytidine Monophosphate / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D008099:Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010802:Phylogeny; D011066:Poly C / Q000378:metabolism*; D011072:Poly U / Q000378:metabolism; D012260:Ribonucleases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D000596:Amino Acids; D011072:Poly U; D011066:Poly C; D012260:Ribonucleases; D003568:Cytidine Monophosphate