Abstract
Onconase, a homolog of ribonuclease A (RNase A) with low ribonucleolytic activity, is cytotoxic and has efficacy as a cancer chemotherapeutic. Here variants of RNase A were used to probe the interplay between ribonucleolytic activity and evasion of the cytosolic ribonuclease inhibitor protein (RI) in the cytotoxicity of ribonucleases. K41R/G88R RNase A is a less active catalyst than G88R RNase A but, surprisingly, is more cytotoxic. Like Onconase, the K41R/G88R variant has a low affinity for RI, which apparently compensates for its low ribonucleolytic activity. In contrast, K41A/G88R RNase A, which has the same affinity for RI as does the K41R/G88R variant, is not cytotoxic. The nontoxic K41A/G88R variant is a much less active catalyst than is the toxic K41R/G88R variant. These data indicate that maintaining sufficient ribonucleolytic activity in the presence of RI is a requirement for a homolog or variant of RNase A to be cytotoxic. This principle can guide the design of new chemotherapeutics based on homologs and variants of RNase A.
Highlights
OnconaseTM, a homolog of ribonuclease A (RNase A) with low ribonucleolytic activity, is cytotoxic and has efficacy as a cancer chemotherapeutic
Like OnconaseTM, the K41R/G88R variant has a low affinity for ribonuclease inhibitor protein (RI), which apparently compensates for its low ribonucleolytic activity
We find that cytotoxicity can be maintained in an RNase A variant with decreased catalytic activity if there is a concomitant decrease in the affinity for RI
Summary
From the ‡Department of Biochemistry and the ʈDepartment of Chemistry, University of Wisconsin, Madison, Wisconsin 53706. OnconaseTM, a homolog of ribonuclease A (RNase A) with low ribonucleolytic activity, is cytotoxic and has efficacy as a cancer chemotherapeutic. Variants of RNase A were used to probe the interplay between ribonucleolytic activity and evasion of the cytosolic ribonuclease inhibitor protein (RI) in the cytotoxicity of ribonucleases. We find that cytotoxicity can be maintained in an RNase A variant with decreased catalytic activity if there is a concomitant decrease in the affinity for RI. This finding has important implications for understanding the biochemical basis for the cytotoxicity of ribonucleases
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