Tryptophan environment in adenosine deaminase. I. Enzyme modification with N-bromosuccinimide in the presence of adenosine and EHNA analogues.
Adenosine deaminase from bovine cerebral hemisphere (white and gray matter) and spleen was treated with N-bromosuccinimide, a reagent known to oxidize selectively tryptophan residues in proteins. Spectrally observable tryptophan modification was accompanied by enzyme inactivation. Tsow graphics revealed that two Trps are essential for the activity of enzyme from both tissues. Enzyme inhibitors and substrate analogues, derivatives of erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA) and adenosine, were able to protect Trp against modification, and this effect correlated in general with the enzyme activity protection. In the presence of adenosine deaza analogues (the noninhibitor tubercidin among them) only two Trps were modified in the fully inactivated enzyme. In the presence of EHNA and its deaza analogues, full inactivation of the enzyme was accompanied by the modification of four Trps. The obtained data confirm the previous hypothesis about the presence on the enzyme of different binding sites for adenosine and EHNA derivatives that are responsible for the different effects on the enzyme conformation elicited by the corresponding derivatives. Moreover, these data allow us to suggest that Trp residues, still unidentified by X-ray analysis, are essential for the functioning of the enzyme.
H. Ch. Buniatyan Institute of Biochemistry of the Armenian NAS, 511 P. Sevak St., Yerevan 375014, Armenia. biochem@ipia.sci.am
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S
Mardanyan
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Sharoyan
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Antonyan
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Armenyan
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Cristalli
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Lupidi
D000225:Adenine / Q000031:analogs & derivatives*; D000241:Adenosine / Q000737:chemistry*; D000243:Adenosine Deaminase / Q000737:chemistry*; D058892:Adenosine Deaminase Inhibitors; D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000201:enzymology; D001974:Bromosuccinimide / Q000737:chemistry*; D002417:Cattle; D004791:Enzyme Inhibitors; D007700:Kinetics; D011487:Protein Conformation; D013154:Spleen / Q000201:enzymology; D014364:Tryptophan / Q000032:analysis / Q000737:chemistry*
D058892:Adenosine Deaminase Inhibitors; D004791:Enzyme Inhibitors; C010192:9-(2-hydroxy-3-nonyl)adenine; D014364:Tryptophan; D000243:Adenosine Deaminase; D000225:Adenine; D000241:Adenosine; D001974:Bromosuccinimide