V.G. Malathi

Effects of murine viral leukemia on spleen nucleoside deaminase: Purification and properties of the enzyme from leukemic spleen☆

Abstract A 400-fold increase was observed in the specific activity of nucleoside deaminase (cytidine aminohydrolase, EC 3.5.4.5) in the spleen of mice infected with Friend murine leukemia virus. The enzyme was purified 100-fold from the spleen of leukemic mice. It was sensitive to SH reagents and stimulated by dithiothreitol and GSH. Different pH responses were observed in the presence or absence of dithiothreitol. Cytidine was deaminated at a greater rate than deoxycytidine; cytosine or its nucleotides were not deaminated. The molecular weight determined by gel filtration was 74 000.

[53] Cytidine deaminase from leukemic mouse spleen

Publisher Summary This chapter describes the purification procedure of cytidine deaminase enzyme from leukemic mouse spleen. Cytidine deaminase is widely distributed among mammalian tissues. This enzyme is responsible for the inactivation of cytosine arabinoside––one of the most useful chemotherapeutic agents available for the treatment of acute myeloblastic leukemia. In the mouse spleen, a marked increase in the specific activity of cytidine deaminase occurs following infection with Friend leukemia virus. Elevated levels of cytidine deaminase are also found in regenerating mouse liver, in human leukemic cells, following the treatment of the patient with cytosine arabinoside, and in HeLa cells cultured in the presence of cytosine arabinoside. The activity of cytidine deaminase is assayed by measuring the amount of labeled uridine formed by the deamination of 2-14C-labeled cytidine. High levels of cytidine deaminase are found in human polymorphonuclear leukocytes.

An Artemia salina factor which counteracts the mRNA-induced inhibition of initiator Met-tRNA binding to initiation factor eIF-2.

Abstract Ternary complex formation between eukaryotic initiation factor 2 (eIF-2), initiator Met-tRNA and guanosine 5′-(β, γ-imino) triphosphate [GMP-P(NH)P] is strongly inhibited by mRNA in the Artemia salina system. Developing A. salina embryos contain a factor which displays a novel activity, namely the ability to counteract the mRNA-induced inhibition of ternary complex formation. This factor is heat-labile. It is proposed that the factor may play an important role in protein biosynthesis by preventing mRNA from inhibiting an early step of peptide chain initiation.

Stimulation of an Artemia salina ATPase activity by eukaryotic messenger RNAs

Summary Several ‘capped’ mRNAs (e.g., 9S globin mRNA) as well as ‘noncapped’ mengo viral mRNA greatly stimulate the activity of a novel ATPase isolated from Artemia salina embryos. Other RNAs tested [tRNAs, initiator Met-tRNA, poly r(U), poly r(A)] are much less stimulatory. The ATPase activity is also stimulated by mRNAs in the presence of 40S ribosomal subunits. However, the other RNAs listed above are inhibitory under the latter conditions. The ATPase activity is strongly stimulated by 40S ribosomal subunits in the absence of mRNAs. It is suggested that the ATPase, described here, might be involved in the binding of natural eukaryotic mRNAs to 40S peptide chain preinitiation complexes, a process known to depend on ATP hydrolysis.

An inhibitor which interferes with the enzymatic aminoacylation of tRNA.

Inactive, frozen and thawed cytoplasmic extracts of 3T3 and SV-101 (3T3 transformed by SV-40 virus) cells contain an inhibitor which blocks the poly(U)-directed incorporation of [14C]phenylalanine into polypeptides, catalyzed by active extracts of these cells. This inhibition is not reversed by adding increased amounts of poly(U). Furthermore, little or no inhibitory activity is observed when poly(U) translation is assayed using precharged [14C]Phe-tRNA. These results suggest that the observed inhibition is not due to the degradation of poly(U) by a nuclease. The inhibitor appears to act primarily at the level of tRNA charging since the synthesis of both Phe-tRNA and Lys-tRNA is impaired in its presence. Evidence is presented which indicates that the inhibitory activity is not due to a high molecular weight protein or nucleic acid. However, the inhibitor appears to be adsorbed to a macromolecule. The inhibitory activity is completely destroyed by ashing.