ABSTRACT
Two thermophilic and thermostable enzymes, isolated from Sulfolobus solfataricus, S-adenosylhomocysteine hydrolase and 5'-methylthioadenosine phosphorylase, were exposed to 10.4 GHz microwave radiation in order to discriminate between thermal and non-thermal microwave effects. The exposure causes a non-thermal, irreversible and time-dependent inactivation of both enzymes; the inactivation rate is related to the energy absorbed and is independent of the enzyme concentration. The influence of salts on enzyme inactivation has also been investigated. Conformational changes of S-adenosylhomocysteine hydrolase, detected by fluorescence and circular dichroism techniques, suggest that microwaves induce protein structural rearrangements not related to temperature.
Subject(s)
Hydrolases/radiation effects , Microwaves , Protein Conformation , Purine-Nucleoside Phosphorylase/radiation effects , Sulfolobus/enzymology , Adenosylhomocysteinase , Circular Dichroism , Enzyme Stability , Hot Temperature , Hydrolases/chemistry , Hydrolases/metabolism , Kinetics , Protein Conformation/radiation effects , Purine-Nucleoside Phosphorylase/chemistry , Purine-Nucleoside Phosphorylase/metabolism , Spectrometry, FluorescenceABSTRACT
We examined the emission spectra of tyrosine- and tryptophan-containing proteins using one-photon (270-310 nm) and two-photon (565-610 nm) excitation. Emission spectra for two-photon excitation of native and denatured human serum albumin and of three purine nucleoside phosphorylases indicated an absence of the tyrosine emission normally seen for one-photon excitation below 290 nm. We examined the one-photon and two-photon excitation spectra of tyrosine-tryptophan mixtures to determine the origin of selective excitation of the tryptophan residues. These results confirmed a short-wavelength shift of the tyrosine two-photon excitation spectrum relative to that of tryptophan, as recently reported by Rehms and Callis (1993) Chem. Phys. Lett. 208, 276-282.
Subject(s)
Proteins/chemistry , Animals , Cattle , Humans , Photochemistry , Photons , Proteins/radiation effects , Purine-Nucleoside Phosphorylase/chemistry , Purine-Nucleoside Phosphorylase/radiation effects , Serum Albumin/chemistry , Serum Albumin/radiation effects , Spectrometry, Fluorescence , Tryptophan/chemistry , Tryptophan/radiation effects , Tyrosine/chemistry , Tyrosine/radiation effectsABSTRACT
Changes in ADA and PNP activities in the spleens and thymuses of mice were studied after a single administration of cyclophosphamide (CY, 200 mg/kg) and after whole-body gamma irradiation (5.5 Gy), applied alone or three days after CY application. In the first days after the treatment the enzyme activities were significantly depressed (p less than 0.01) with the exception of ADA in the spleen, where a high elevation (220-380%) in relation to controls was observed. During the regeneration period a pronounced rise of PNP activity in the spleen occurred mainly after a combined application of CY and irradiation (270%). In the thymus the regeneration was manifested by a mild increase of both ADA and PNP activities towards control values. The findings suggest that the expressive changes of ADA and PNP activities, participating in the purine salvage pathway, may, after a cytotoxic treatment, influence the nucleotide pool and DNA synthesis in lymphoid organs.
Subject(s)
Adenosine Deaminase/radiation effects , Cyclophosphamide/pharmacology , Purine-Nucleoside Phosphorylase/radiation effects , Spleen/drug effects , Thymus Gland/drug effects , Adenosine Deaminase/metabolism , Animals , Chromatography, High Pressure Liquid , Gamma Rays , Injections, Intraperitoneal , Mice , Mice, Inbred C57BL , Mice, Inbred CBA , Organ Size , Purine-Nucleoside Phosphorylase/metabolism , Spleen/enzymology , Spleen/radiation effects , Thymus Gland/enzymology , Thymus Gland/radiation effects , Whole-Body IrradiationABSTRACT
The activities of adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP) were determined between days 1-14 in the spleen, thymus and femoral bone marrow of mice subjected to whole-body gama irradiation with a dose of 5.5 Gy. In control animals, the highest activity of ADA (as related to 10(6) cells) was recorded in the thymus (58.9 pmol.s-1), the lowest one in the femur (34.8 pmol.s-1), the PNP activity was the lowest in the thymus (14.5 pmol.s-1) and the highest in the femur (96.0 pmol.s-1). In the spleen, an elevation of ADA activity (up to 379%) was observed during the first postirradiation days; PNP activity was reduced (to 58%) on postirradiation day 3, followed by the return and even elevation on day 14 (265%). In the thymus, a parallel reduction of the activities of both enzymes appeared during the first postirradiation days, with a subsequent increase during the regeneration phase. In the femoral bone marrow, ADA and PNP activities were increased on postirradiation day 1 (275% and 201%, respectively). Reference is made to the possible relationship between the observed characteristic changes in activities and the degree of damage and/or renewal of cell population in the hemopoietic tissues after irradiation.
