[Blood cobalamins and xenobiotic-metabolizing enzymes in rat liver in adjuvant arthritis]. / Kobalaminy krovi i fermenty metabolizma ksenobiotikov v pecheni krys pri ad''iuvantnom artrite.

Changes in the total cobalamin content and spectrum of individual forms of these vitamins in blood cells and plasma as well as the activities of enzymatic systems of xenobiotic metabolism in liver microsomes of rats with experimental adjuvant arthritis (AA) have been studied. The total cobalamin content in the blood plasma of rats with AA was increased in comparison with intact animals; however, leucocytes from AA rats were deficient in methylcobalamin (MeCbl). A correlation was found between the ratios of individual cobalamin forms and their total content which was differently expressed in experimental and control animals. The development of AA was associated with marked inhibition of the cytochrome P-450-dependent monooxygenase system of the liver and glutathione transferase. The possibility of correction of these disturbances by MeCbl is discussed.

[Effect of various factors on the methylation of DNAse I]. / Vliianie nekotorykh faktorov na reaktsiiu metilirovaniia DNKazy I.

The effects of NaCl, EDTA and tRNA on methylation and enzymatic properties of deoxyribonuclease I (DNase I) were investigated. The methylation was carried out by S-methylmethionine (vitamin U) in the phosphate-citric buffer pH 4.0. It was found that 0.5 M NaCl decreases by about 30% the incorporation of CH3-groups into the DNase, whereas 1.5 M NaCl-by 47%. A similar, although a less pronounced effect, was exerted by tRNA within the concentration range of 1.36-34.7 microM. On the contrast, EDTA (0.01-0.05 M) stimulated the incorporation of CH3-groups by 15 and 30%, respectively. The functional properties of methylated DNase I in the presence of EDTA remained unaffected. The enzyme methylation in the presence of NaCl or tRNA caused deceleration of the 3H-DNA hydrolysis (by 15-30%) only within the first 20 min of the reaction.

[Effect of 5-azacytidine on E. coli cells with different DNA-methylases]. / Deistvie 5-azatsitidina na kletki E. coli s razlichnymi DNK-metilazami.

A correlation was found between the bacteriocide effect of 5-aza-C and the amount of cytosine DNA-methylases in E. coli cells. 5-Aza-C-DNA induced partial or complete inhibition of bacterial DNA-methylases with different site specificity; cytosine DNA-methylases were inhibited by the DNA more effectively than adenine DNA-methylase Eco dam. The inhibitory influence of 5-aza-C-DNA on cytosine DNA-methylases was due to the formation of stable inactive complexes between the enzyme and the non-methylating cytosine analog in the recognition sites. Cytosine DNA-methylase Eco RII formed a relatively firm bond with 5-aza-C-DNA, which could be disrupted by 1 M KCl; this disruption restores the DNA-methylase activity and the inhibiting capacity of 5-aza-C-DNA. Thus, the binding of cytosine DNA-methylase to 5-aza-C in DNA is noncovalent; the inhibition of the enzyme by 5-aza-C-DNA is reversible.

[DNA methylation in the cells of Escherichia coli MRE 600 in the presence of S-methylmethionine]. / Metilirovanie DNK v kletkakh Escherichia coli MRE 600 v prisutstvii S-metilmetionina.

The effect of S-methylmethionine (SMM), a methyl group donor, on enzymatic methylation of DNA in E. coli MRE 600 cells was studied. It was found that SMM can be used as a donor of methyl groups during bacterial DNA methylation in vivo without changing the specificity of DNA methylation.

[Cobalamins and tRNA methyltransferase activity in E. coli cells]. / Kobalaminy i tRNK-metiltransferaznaia aktovnost'kletok E. coli.

The end of the logarithmic phase of growth of E. coli 113-3 cells in a mineral medium with methylcobalamin (MeCbl) occurs 60 minutes earlier than in the case of cells grown with cyancobalamin (CNCbl). Chromatography on a Sephadex - DEAE cellulose column of the 105 000xg fractions of cell-free extracts of the cells grown in the presence of MeCbl showed both quantitative and qualitative differences. The ability of individual protein fractions to transfer the 14CH3-groups from the Adomet to submethylated tRNA of E. coli K12W6 in the cells grown with MeCbl increases 1.5 - 3-fold in comparison with those grown with CNCbl. A chromatographic analysis of methylated bases of tRNA did not show any qualitative differences. In the presence of MeCbl the incorporation of the CH3-groups into m7G is increased by 50%.

[Vitamin U and RNA metabolism in prokaryotes]. / Vitamin U i obmen RNK u prokariot.

The paper is concerned with a study of the vitamin U effect on the rate of 14C-uridine incorporation into various categories of RNA in E. coli MRE-600 cells. It is found that cells grown with vitamin U (0.06 mg/ml) and incubated with 14C-uridine for 5 min are able to produce a 10-12-fold increase of the label incorporation into 4 S and 5 S RNA and a 14-fold increase into high polymeric RNA in comparison with the control cells. Under longer intervals of incubation (20 min) the intensity of high-polymeric RNA formation was half as high as for 4 S and 5 S RNA formation. MAK column chromatography of high-polymeric RNA in salt and temperature gradients showed the presence of the RNA temperature fraction in bacteria cells. Vitamin U stimulates the formation of various categories of RNA and causes a quantitative increase in the RNA temperature fraction.

[Influence of S-methylmethionine on the metabolism of nucleic acids in Escherichia coli]. / Vliianie S-metilmetionine na metabolizm nukleinovykh kislot y E. coli.

The influence of S-methylmethionine (SMM) on metabolism of nucleic acids in E. coli MRE-600 has been studied. The influence of SMM was compared with that of methionine. Addition of SMM to the medium at the concentration of 2.7 . 10(-5) M increased the incorporation of 5-methyl-3H-thymidine into DNA by about 20% and the incorporation of 2-14C-uridine into the total RNA of the cells by about 47%. The electrophoretic separation of RNA in polyacrylamide gels showed that SMM stimulated the incorporation of the label into 4 S, 5 S, 16 S and 23 S RNA with different intensity. The highest stimulation of the label incorporation has been observed in 16 S RNA.

[Methylcobalamin-dependent methylation of proteins during malignant degeneration]. / Metilkobalaminzavisimoe metilitrovanie belkov pri malignizatsii.

A capacity of methylcobalamine (14CH3-B12) to methylate proteins from rat liver tissue and from Zajdela ascites hepatoma was studied. The rate of methylation using 14CH3B12 was compared with that of the universal donor of methyl groups--S-adenosyl-L-(methyl-14C[methionine], Ado-met)-14CH3-B12 methylated proteins from Zajdela ascites hepatoma 6-9-fold and proteins from liver tissue 50-60-fold more intensively as compared with Ado-met in vitro. The methylating ability of 14CH3-B12 depended on protein concentration. Liver proteins were methylated at the rate 2-2.5-fold higher than Zajdela ascites hepatoma proteins. At the same time, Ado-met was the most effective donor of 14CH3 groups for proteins of Zajdela ascites hepatoma. Heat treatment of proteins increased the rate of their methylation using 14CH2-B12 by 10-20%; when Ado-met was used the transfer of CH3 groups was decreased 2-5-fold.

[tRNA methyltransferase activity in Escherichia coli growing on a medium with methionine and S-methylmethionine]. / Metiltrsanferaznaia aktivnost' tRNK u Escherichia coli pri ee rosta na srede s metioninom i S-metilmetioninom.

tRNA methylating enzymes isolated from E. coli 113-3 grown in a medium with methionine or S-methylmethionine differed in the activity of transfer of the 14CH3-group from 14CH3-S-adenosyl-L-methionine (SAM) to tRNA in the in vitro experiments in heterological systems consisting of tRNA methyltransferases from E. coli 113-3 and a substrate of methylation, viz. methyl-deficient tRNA isolated from E. coli K 12W6. In such a system, tRNA methyltransferases from E. coli 113-3 grown on S-methylmethionine methylated tRNA from E. coli K 12W6 at a rate almost twice as high as that in the case of the culture grown on methionine. At the same time, the activity of guanine methylation yielding 7-methylguanine increased significantly. The protein obtained after precipitation of cell-free extracts with ammonium sulfate was separated into four fractions using a system of columns packed with Sephadex G-25 and DEAE-cellulose. These fractions differed quantitatively in culture grown either on methionine or S-methylmethionine.

[Methylation of ribonuclease and albumin by methyl cobalamin]. / Metilirovanie ribonukleazy i al'bumina metilkobalaminom.

The capacity of methyl cobalamine (14CH3-B12) to methylate RNase and albumin in in vitro systems was studied. Under the experimental conditions, 14CH3-B12 methylated proteins about 100--1000 times more actively than S-adenosyl-methionine, a universal donor of methyl groups. The nature of buffer and pH 2 to 8 had no noticeable effect on the methylating capacity of 14CH3-B12. However, at higher pH rate of incorporation of methyl groups slightly increased. The 14CH3-groups incorporated into RNase amino acids remained stable upon illumination, in the presence of KCN in the incubation medium, and when heated in 20% HCl for 24--72 hr at 105 degrees. Methylation did not influence the enzymic properties of RNase. The automatic amino acid analyzer showed three peaks of the label of modified amino acids in the hydrolzates of methylated RNase, one of which corresponded to methylated methionine.

[Methylation of prokaryotic RNA by S-methyl methionine in in vivo experiments]. / Metilirovanie prokarioticheskikh RNK S-metilmetioninom.

While cultivating the E. coli 113-3 strain on the mineral medium containing S-methyl-(methyl-3H)-methionine, the incorporation of methyl groups into 4S, 16S and 23S RNA proved to be over 5 times more effective as compared with the control, when L-(methyl-3H)-methionine acted as a donor of methyl groups. The ratio of methylated components has much in common and significant differences. All the types of RNA of E. coli grown in the mineral medium containing S-methyl methionine showed an increased ratio of m2G as compared with the corresponding types of RNA of E. coli after its cultivation in the methionine containing mineral medium.

[Some aspects of protein methylation]. / Nekotorye aspekty metilirovaniia belka.

The paper gives a review of the literature publications on protein methylation and probable donors of methyl groups. It presents experimental data demonstrating the capability of 14CH3-B12 to methylate in vitro proteins of tRNA methylases from Zajdela ascite hepatoma and rat liver as well as commercial RNase and albumin preparations.

[Influence of methylcobalamin and cyanocobalamin on the neoplastic process in rats].

The effect of methylcobalamine (5.6-dimethylbenzimidazolyl-Co-methylcobamide, CH3-B12) and cyanocobalamine (5,6-dimethylbenzimidazolyly-Co-cyanocobamide, CN-B12) on the growth of Walker's carcinosarcoma and longevity of white noninbred rats with implanted Zajdela ascites hepatoma was studied. The two agents exerted a similar effect. They 1) reduced the survival of rats with implanted Zajdela ascites hepatoma and Walker's carcinosarcoma; 2) did not increase the cell concentration in ascites; and 3) increased the total volume of ascites.

[Effect of cyanocobalamin, cobamide coenzyme and the cytostatic hispen on changes in cobalamin levels in healthy rats and rats with transplanted sarcoma M-1]. / Vliianie tsiankobalamina, kobamidnogo kofermenta i tsitostatika gisfena na izmenenie urovnia kobalaminov u zdorovykh krys i krys s privitoi sarkomoi M-1

The influence of cyanocobalamine (CN-B12), cobamide coenzyme (5'-deoxyadenosyl-B12, DBC) and the cytostatic hisphen on the level of cobalamines in the blood serum, liver, kidney, heart and in the tumor of healthy and sarcoma M-1 implanted rats was studied. In the above organs and tissues cobalamines were found to occur in the free and protein-bound state (cobalamine-protein complexes). The complexes were of different stability. The ratio of free to protein-bound cobalamines varied. The content of cobalamine-protein complexes in healthy and tumorous tissues was different. CN-B12, DBC and hisphen influenced specifically the redistribution of cobalamine and protein-bound complexes.

[Cobalamin levels in rats with transplanted Zajdela ascites hepatoma]. / Uroven' kobalaminov u krys s privitoi astsitnoi gepatomoi Zaidelia

The level of cobalamines in the kidney and liver of healthy rats and rats with implanted Zajdela ascites hepatoma (ZAH) was examined. The results were compared with the data obtained from the rats with sarcoma M-1. It was shown that different tumors had dissimilar effects upon the level of total cobalamines. However, the ratio of free cobalamines to protein-bound cobalamines (cobalamine-protein complexes) varied. The concentration of cobalamine-protein complexes increased in the liver, kidney and in the tumor. The experiments with ZAH demonstrated that ZAH cells contained appreciable amounts of cobalamines, whereas the supernatant contained only a minor quantity of cobalamines. Injections of 5'-deoxyadenosyl-B12 increased the level of cobalamines in ZAH cells and had a low effect on the level of cobalamines in the supernatant.

[Effect of cobamide coenzyme on the growth of several transplantable tumors in rats and on prolonging the life of rats with grafted Zajdel ascitic tumor]. / Vliianie kobamidnogo kofermenta na rost nekotorykh privivaemykh opukholei u krys i na prodlenie zhizni krys s privitoi astsitnoi geptaomoi Zaidelia

The effect of the cobamide coenzyme (5,6-dimethylbenzimidazolyl-Co-5'-deoxyadenozylcobamide, DBC) on the growth of some forms of rat tumours and on the prolongation of survival of rats with implanted Zajdela ascites hepatoma was compared with that of cyanocobalamine (5,6-dimethylbenzimidazolyl-Co-cyanocobamide, CN-B12). The effect of DBC was shown to differ from that of CN-B12. DBC did not stimulate the growth of the investigated forms of tumours and prolonged survival of rats with implanted Zajdela ascites hepatoma. The rats displayed cancer resistance when DBC or CN-B12 were injected before implantation of tumours or Zajdela ascites hepatoma.

[Participation of 5'-deoxyadenosyl-B12 in regulating methylation of tRNA]. / Uchastie 5'-dezoksiadenozil-B12 v reguliatsii metilirovaniia tRNK

The effects of different forms of cobalamines on the activities of tRNA-methylases of Zajdela ascite hepatoma were studied. Of six cobamides studied 5'-deoxyadenosyl-B12 and factor B containing as a ligand HSO3 in the concentrations of 2.4-10(-5) and 4.8-10(-5) M inhibited the tRNA-methylase activity by 21% and 15% correspondingly. The inhibitory effect of 5'-deoxyadenosyl-B12 is probably dependent on the adenosyl part of the molecule. 5'deoxyadenosyl-B12 exerted a selective effect of Zajdela ascite hepatoma tRNA-methylases, inhibiting largely the activity of 5-methyl cytosine methylase during the methylation of the E. coli K12W6 tRNA and yeast tRNA1 Val.

[Estimation of kinetic constants and study of site specificity of Zajdela ascite hepatoma and rat liver tRNA-methylases]. / Opredelenie kineticheskikh konstant i izuchenie pozitsionnoi spetsifichnosti tRNK-metilaz astsitnoi gepatomy zaidelia i pecheni krys

Individual yeast tRNAVal1 was used as a substrate for estimation of kinetic constants and study of site specificity of m5C-and m1A-methylases of Zajdela ascite hepatoma and rat liver. It was demonstrated that the rate of yeast tRNAVal1 methylation by hepatoma tRNA-methylases is 4--5 times higher than that induced by liver tRNA-methylases. The rates of 1-hour methyl groups incorporation into tRNAVal1 were 3.7 and 4.7 times higher in case of m5C-and m1A-methylases and 9.4 and 4.5 times higher in case of m1G-and m7G-methylases of hepatoma than the respective rates obtained for corresponding liver methylases. The main products of methylation were m5C and m1A containing about 90% of total radioactivity incorporated into tRNA. m5C-methylases of liver and hepatoma had similar affinity for S-Ad-Met. The Km value for both enzymes was 2.66 micronmole; the Km values for m1A-methylases of liver and hepatoma with respect to S-Ad-Met were the same and equal to 0,25 micronmole. m5C and m1A methylases of liver and hepatoma had adequate affinity for yeast tRNAVal1; their site specificity was the same, since they methylated in yeast tRNAVal1 cytosine in the tetracytidylic sequence of C49--C52 and adenine in the 59th position from the 5'-end of the molecule.