[Conformation of adenosine deaminase in complexes with inhibitors: application of selective quenching of fluorescence emission].

The effect of inhibitors, 1-deazaadenosine (1-dAdo) and erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), on the conformation of adenosine deaminase was studied using the method of selective quenching of fluorescence emission by acrylamide, I- and Cs+. Both in free adenosine deaminase and in its complexes with the inhibitors, the wavelength maxima and half-width of the emission characterize the environment of fluorescing tryptophan residues in adenosine deaminase as weak polar with limited access to solvent. The formation of complexes with the ground state inhibitors used did not quench or change the main emission characteristics of tryptophan fluorescence in adenosine deaminase. Small blue shifts of emission maxima were observed upon quenching in all three samples. The Stern-Volmer parameters of tryptophan fluorescence quenching by acrylamide were not essentially influenced by complex formation of the enzyme with the inhibitors: in general, the folding of the enzyme molecule in the complexes is not perturbed. On the contrary, the emission quenching by charged heavy ions, I- and Cs+, in the complexes was hindered in comparison with free adenosine deaminase. In the complex with 1-deazaadenosine, the parameters for quenching by both ions evidence the essential worsening of their interaction with tryptophans. In the complex with erythro-9-(2-hydroxy-3-nonyl)adenine, along with the worse quenching by I-, complete prohibition of quenching by Cs+ was observed. These data indicate that the local environments of fluorescing tryptophan residues is substantially distorted compared with free adenosine deaminase, which leads to their screening from charged heavy ions.

[Activity of pleural fluid adenosine deaminase in tuberculous pleurisy]. / Aktivnost' adenozindezaminazy plevral'noi zhidkosti pri tuberkuleznom plevrite.

Examining the activity of adenosine deaminase in the pleural fluids of 69 patients with tuberculous pleurisy of various etiology from the clinics of Armenia indicated that it was greater than the threshold value of 20 U/L in 95.7 of 47 patients with tuberculous pleurisy. The specificity of this parameter for this disease was 0.91. The prognostic value of the test with positive and negative results was 0.96 and 0.94, respectively. The diagnostic value of the ADA test was 0.94.

[The role of tryptophan in appearance of adenosine deaminase activity]. / O roli triptofana v proiavlenii aktivnosti adenozindezaminazy.

Chemical modification of tryptophan residues with N-bromosuccinimide and their photooxidation in the presence of trichloroethanol inhibited the activity of adenosine deaminase purified from gray and white matter of calf brain. Only two of six modified residues are important for enzyme activity. Preliminary kinetic data indicate that these essential tryptophan residues are adjacent to the substrate-binding site of the enzyme.

[Isolation, purification, and comparative study of the properties of adenosine deaminase from five regions of the cattle brain]. / Vydelenie, ochistka i sravnitel'noe izuchenie svoistv adenozindezaminazy iz piati otdelov mozga krupnogo rogatogo skota.

Adenosine deaminase from the white and gray matter of the large hemispheres, cerebellum, medulla oblongata and pituitary anterior lobe has been isolated and purified. The pH optimum, Km, molecular mass, yield and specific activities for all the enzyme preparations have been determined. Gel filtration and electrophoresis data point to the heterogeneity of the enzyme. The lack of effects of SH-reagents suggests the absence of an essential SH-group. Among five bivalent metal ions, only Cu2+ irreversibly inhibited the enzyme activity in all the preparations.

The role of tryptophanyl residues in electron transfer from NADPH--adrenodoxin reductase to adrenodoxin.

Chemical modification of tryptophanyl residues of NADPH - adrenodoxin reductase by N - bromosuccinimide and trichloroethanol prevents the interaction of the enzyme with adrenodoxin. The modification does not touch other amino acid residues besides tryptophan (tyrosine, lysine and cysteine) or disturb the structure of protein. The presence of adrenodoxin suppresses the modification. The data obtained indicate the participation of adrenodoxin reductase tryptophan residues in the interaction with adrenodoxin.

[Subunit structure of AMP-deaminase from skeletal muscles]. / Sub''edinichnoe stroenie AMP-dezaminazy skeletnykh myshts.

AMP-deaminase was purified from skeletal muscle of rat by the affinity chromatography on phosphocellulose and gel-filtration on Sephadex G-200. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate (SDS-PAGE) has shown three protein bands on each step of purification. One of them corresponds to the subunit of tetrameric AMP-deaminase molecule with molecular weight of 76 kDa and two others--to the protein subunit with molecular weight of 42 and 33 kDa. Repeated SDS-PAGE of the main subunit band has revealed again all these protein bands. The data obtained indicate that AMP-deaminase subunit of 76 kDa is able to dissociate on two polypeptide chains with similar values of molecular weights in the presence of SDS.