Extending the limits to enzymatic catalysis: diffusion of ribonuclease A in one dimension.
Bovine pancreatic ribonuclease A (RNase A) is a distributive endoribonuclease that catalyzes the cleavage of the P-O5' bond of RNA on the 3' side of pyrimidine residues. Here, RNase A is shown to cleave the P-O5' bond of a pyrimidine ribonucleotide faster when the substrate is embedded within a longer tract of poly(adenylic acid) [poly(A)] or poly(deoxyadenylic acid) [poly(dA)]. These data indicate that a ribonuclease can diffuse in one dimension along a single-stranded nucleic acid. This facilitated diffusion is mediated by Coulombic interactions, as the extent is diminished by the addition of NaCl. RNase A is more effective at cleaving a pyrimidine ribonucleotide embedded within a poly(dA) tract than within a poly(deoxycytidylic acid) [poly(dC)] tract. T45G RNase A, which catalyzes the processive cleavage of poly(A) but the distributive cleavage of poly(cytidylic acid) [poly(C)], has the same preference. Apparently, processive catalysis by the T45G enzyme arises from the expanded substrate specificity of the variant superimposed upon an intrinsic ability to diffuse along poly(A). Homologous ribonucleases with cytotoxic activity may rely on facilitated diffusion along poly(A) tails for efficient degradation of the essential information encoded by cellular mRNA.
Kelemen
B R
BR
Department of Biochemistry, Department of Chemistry, University of Wisconsin-Madison 53706, USA.
Raines
R T
RT
D000818:Animals; D002384:Catalysis; D002417:Cattle; D004058:Diffusion; D005452:Fluoresceins / Q000378:metabolism; D006147:Guanine / Q000378:metabolism; D006868:Hydrolysis; D015345:Oligonucleotide Probes / Q000138:chemical synthesis / Q000235:genetics; D011061:Poly A / Q000378:metabolism; D011066:Poly C / Q000378:metabolism; D012259:Ribonuclease, Pancreatic / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity / Q000235:genetics; D013941:Thymine / Q000378:metabolism
D005452:Fluoresceins; D015345:Oligonucleotide Probes; D011061:Poly A; C015465:poly(dA); C023826:poly(dC); D011066:Poly C; D006147:Guanine; D012259:Ribonuclease, Pancreatic; D013941:Thymine