Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.27.3 (RNase T1)
 1,228 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The single tryptophan residue in ribonuclease T1 [EC 3.1.4.8] was selectively oxidized by ozone to N'-formylkynurenine, which was then converted to kynurenine by acid-catalyzed deformylation in the frozen state. The two enzyme derivatives thus formed, NFK- and Kyn-RNase T1, lost enzymatic activity at pH 7.5, at which native RNase T1 most efficiently catalyzes the hydrolysis of RNA. At pH 4.75, the modified enzymes retained a decreased but distinct enzymatic activity toward RNA without alteration of substrate specificity, and Kyn-RNase T1 was four times more active than NFK-RNase T1. The binding of 3'-GMP to these modified enzymes decreased remarkably at pH 5.5, the optimum pH for binding to the intact enzyme. The gamma-carboxyl group of glutamic acid 58 was still reactive to iodoacetic acid after modification of tryptophan 59. The amounts of the carboxymethyl group introduced into NFK- and Kyn-RNase T1 were 0.36 and 0.59 mol, respectively, under conditions such that quantitative esterification of native RNase T1 takes place. CD spectroscopy indicated that the tertiary structure of the molecule was disordered in NFK-RNase T1, but not significantly in Kyn-RNase T1. It is concluded that tryptophan 59 functions in maintaining the active conformation of the protein structure, particularly in constructing the active environment for a functionally important set of groups involved in the binding of the substrate at the active site, although direct participation of in tryptophan the catalytic function of ribonuclease T1 is unlikely.
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PMID:Chemical modification of ribonuclease T1 with ozone. 41 75

Ribonuclease T1 (RNase T1) carboxymethylated at the gamma-carboxyl group of Glu-58 with iodoacetic acid is known to be completely inactive while it retains an almost full substrate-binding ability. In order to further clarify the effects of the carboxymethylation, the thermal stabilities of intact and Glu-58-carboxymethylated (CM-) RNase T1 were compared by measuring 1H NMR spectra at various temperatures. The transition curves of unfolding were obtained by plotting, as a function of temperature, the peak areas for the alpha and delta protons of Asn-81 and Ile-90, respectively, which are well apart from each other in the three-dimensional structure of the enzyme. For each of intact and CM-RNase T1, the transition curve of the Asn-81 alpha proton was identical with that of the Ile-90 delta methyl protons, suggesting that the thermal unfolding occurred simultaneously in every part of the molecule of CM-RNase T1 as well as of intact RNase T1. The midpoint of unfolding was 52 degrees C for intact RNase T1, and was increased by 9 degrees C upon carboxymethylation at Glu-58. This marked stabilization by carboxymethylation is thought to be due to formation of a salt bridge between the introduced carboxymethyl group and the neighboring guanidium group of Arg-77.
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PMID:Thermal stabilization of ribonuclease T1 by carboxymethylation at Glu-58 as revealed by 1H nuclear magnetic resonance spectroscopy. 791 96