In Vitro Incorporation of Selenomethionine into Protein by Vigna radiata Polysomes

Abstract

Vigna radiata polysomes efficiently incorporated 175Selselenomethio-nine, I"4Clmethionine, and I'4Clleucine in vitro. The optimal conditions for translation were determined to be 4.8 millimolar Mg2e, 182 milHimolar K+, and pH 7.4. The rates of incorporation of 175Selselenomethionine and 1'4CImethionine were similar when measured separately, but 175Selseleno-miethionine incorporation was 35% less than 14Clmethionine incorporation when both amino acids were present in equal molar concentrations. Polyacrylamide gel electrophoresis of the hot trichloroacetic acid precipitable translation products demonstrated synthesis of high molecular weight labeled proteins in the presence of 175Selselenomethionine or I35SImethi-onine. No major differences in molecular weights could be detected in the electrophoretic profiles. Utilization of selenomethionine during translation by Vigna radiata polysomes establishes a route for the assimilation of selenomethionine by plants susceptible to selenium toxicity. alteration of this type could give rise to abnormal properties such as altered tertiary structure or abnormal catalytic activity in newly synthesized proteins. As an example of such alterations, the Escherichia coli fi-galactosidase was found to withstand some substitution of methionine by selenomethionine, but an inactive 4S protein with high amounts of substitution was also generated (10). The 4S protein was interpreted to be an inactive subunit of ,Bgalactosidase, produced in excess.Though selenium toxicity to plant systems has been extensively studied with intact plants (23), an understanding at the subcellular level has been limited by the dearth of studies in this area of selenium research. To provide evidence for the in vitro incorporation of selenomethionine into protein, Vigna radiata polysomes and wheat germ supematants were prepared, and the selenium containing proteins synthesized in vitro were analyzed. Our results showed that the [75Se]selenomet2 labeling pattem differed in some respects from ["CJmet incorporation, yet [ 5Selselenomet was rapidly incorporated into high mol wt proteins.Selenium and sulfur constitute an isosteric pair based on chemical similarities and, therefore, have the potential to act as an analog pair in biological processes. Such an analog relationship was evinced from studies in which pasture plants, fed selenite, were able to synthesize the selenoanalogs of several typical sulfurcontaining metabolites (21). Other nonaccumulator plants, such as bromegrass (13) and duck week (21), also synthesize selenomethionine, an analog of methionine, when exposed to selenite. Selenomethionine synthesis in plants may protentiate a toxicity syndrome through indiscriminate utilization of the analog in biochemical reactions which normally use methionine.Methionine is a crucial compound in a multitude of metabolic processes. During the translation process, methionine participates in two key reactions, initiation of protein synthesis and internal insertion of methionine into proteins. In eukaryotes, th...

Differences in Selenium Uptake, Distribution and Expression of Selenium Metabolism Genes in Tomatoes

“…There is a great difference in the plant uptake, transformation and distribution of Se (VI) and Se (IV), and Se (VI) and Se (IV) absorption and transport mechanism are relatively independent. Eustice et al (1980) found that selenate was more easily transported than selenite and the organic Se selenomethionine (SMT), Terry et al (2000) demonstrated the same conclusion. When selenate was absorbed, the ratio of plant Se concentration and root Se concentration was 1.4-1.72, while the ratio range was 0.6-1 when they absorbed selenomethionine (SMT); the ratio range was <0.5 when they absorbed selenite (Zayed et al, 1998).…”

Selenium Toxicity: Aminoacylation and Peptide Bond Formation with Selenomethionine

“…A few comparable in vitro plant studies have focused on the following three aspects of selenium biochemistry: (a) aminoacylation of tRNA with selenoamino acids (2,3,21,24); (b) interactions of selenomethionine with methionine adenosyltransferase (12); and (c) the translational incorporation of selenomethionine into protein (5). Yet, in none of these studies in which selenium-sensitive plants were used, was a biochemical reaction correlated with selenium toxicity.…”

“…One of these analogs, selenomethionine, is known to be formed by a wide variety of plants (7,9,(16)(17)(18). Although the biochemical characteristics of selenomethionine resemble those of methionine in a great many ways, the metabolic activity of selenomethionine apparently differs in certain respects; one of these differences is, in general, reaction rate (4,5,11,12). Examination of those reactions in which selenomethionine functions at a rate different than methionine could reveal some of the factors that underlie selenium toxicity.…”

“…Alteration of initiation process of translation or with the chain elongation process could drastically affect normal functions of the cell. Previous in vitro studies with polysomes from Vigna radiata root tips have established that methionine is a slightly better substrate than is selenomethionine during chain elongation, provided that both amino acids are supplied simultaneously in equal molar concentrations (5).…”

In Vitro Incorporation of Selenomethionine into Protein by Astragalus Polysomes

“…Preparation of wheat germ postribosomal supernatant solutions (S-150) and methods for the translation assays were carried out as described previously (1).…”