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Biomedical subjects

T D Kennedy

Publications and source records attributed to T D Kennedy.

33 records · Page 2Linked to original sources
PubMed1982

Synthesis and turnover of proteins and mRNA in germinating wheat embryos.

The most prominent methionine-labeled protein made when cell-free systems are programmed with bulk mRNA from dry wheat embryos has been identified with what may be the most abundant protein in dry wheat embryos. The protein has been brought to purity and has a distinctive amino acid composition, Gly and Glx accounting for almost 40% of the total amino acids. Designated E because of its conspicuous association with early inhibition of dry wheat embryos, the protein and its mRNA are abundant during the "early" phase (0--1 h) of postimbibition development, and easily detected during "lag" phase (1--5 h), but they are almost totally degraded soon after entry into the "growth" phase of development, by about 10 h postimbibition. The most prominent methionine-labeled protein peculiar to the cell-free translational capacity of bulk mRNA from "growth" phase embryos is not detected as a product of in vivo synthesis. Its electrophoretic properties and its time course of emergence, after 5 h postimbibition development, suggest that this major product of cell-free synthesis may be an in vitro counterpart to a prominent methionine-labeled protein made only in vivo, by "growth" phase embryos. Designated G because of its conspicuous association with "growth" phase development, the cell-free product does not comigrate with any prominent dye-stained band in electrophoretic distributions of wheat proteins. The suspected cellular counterpart to G, also, does not comigrate with a prominent dye-stained wheat protein during electrophoresis, and although found in particulate as well as soluble fractions of wheat embryo homogenates it is not concentrated in either nuclei or mitochondria, as isolated.

Amino Acids↗
PubMed1981

Structural integrity of DNA and translational integrity of ribosomes in nuclease-treated cell-free protein synthesizing systems prepared from wheat germ and rabbit reticulocytes.

After treatment at a microsomal nuclease concentration too low to reduce the endogenous amino acid-incorporating activity of freshly prepared reticulocyte lysate, there is little, if any, intact 26 S RNA left in the ribosomes of either wheat germ or rabbit reticulocyte cell-free protein synthesizing extracts. The primary scissions, probably at highly exposed sites in the rRNA of plant and animal ribosomes, produce two fragments which remain complexed until thermal denaturation reveals "hidden breaks." Molecular weights of the fragments are approximately 0.5 x 10(6) and 0.8 x 10(6) in the case of wheat, and 0.4 x 10(6) and 1.3 x 10(6) in the case of rabbit. There is little perceptible degradation of 5 S, 5.8 S, and 18 S rRNA, or of tRNA in the same extracts. Even though limited degradation of 26 S rRNA by a reticulocyte nuclease has been reported to severely impair the translational mechanism in reticulocyte ribosomes, micrococcal nuclease-induced degradation of rRNA, whether limited or extensive, does not seriously impair the ability of reticulocyte lysates to discriminate, by selective polypeptide synthesis, between complex populations of cellular mRNA. In an allied study, it is shown that under conditions well suited to recovery of the 5.8 S/26 S rRNA complex, with its naturally occurring hidden break, 5 S/18 S rRNA complexing is not detectable in the RNA or metabolizing embryos, nor in the RNA from untreated or nuclease-treated protein synthesizing extracts from wheat and rabbit. The significance of this finding is briefly elaborated in relation to the suggestion that 5 S rRNA may interact with the M2(6)A-m2(6)A hairpin near the 3'-end of 18 S rRNA.

Animals↗
PubMed1979

Wheat embryo ribonucleates. XIV. Mass isolation of mRNA from wheat germ and comparison of its translational capacity with that of mRNA from imbibing wheat embryos.

Commercially milled wheat germ is shown to be a convenient source material for facile recovery of mass (milligram) quantities of highly purified poly(A)-rich RNA. This poly(A)-rich RNA is efficiently translated in a nuclease-treated extract of rabbit reticulocytes. By sucrose density gradient fractionation of bulk poly(A)-rich RNA from wheat germ, it has been possible to show that there is a direct relationship between the molecular weights of the polypeptide products of cell-free synthesis and the molecular weights of the wheat mRNA molecules which program their synthesis. As assessed by SDS -- polyacrylamide gel electrophoresis, the same array of polypeptides is synthesized when nuclease-treated reticulocyte extract is programmed by poly(A)-rich RNA from either commercially supplied or laboratory-prepared wheat embryos. Significantly, there are gross quantitative if not qualitative differences between the translational capacities of poly(A)-rich RNA from dry and imbibing wheat embryos, and the possible importance of these differences for interpreting a changing pattern of polypeptide synthesis in imbibing wheat embryos is the subject of a brief discussion.

Animals↗
PubMed1977

Wheat embryo ribonucleates. IX. Generation of N2-dimethylguanylate when bulk wheat embryo tRNA is used as substrate for wheat embryo S-adenosylmethionine-tRNA methyltransferases, in vitro.

Although the cellular ribonucleates in normally growing cells are virtually saturated with respect to their customary complement of methyl substituents, it has often been reported that 'marginal' levels of (homologous) methylation can be detected when ribonucleates and enzymes from the same source material are incubated, together with S-adenosylmethionine, in vitro. Experiments were designed to acquire new insights that might be useful for circumscribing the number of possible interpretations that could be advanced to account for the introduction of 'supernumerary' methyl groups during (homologous) methylation of wheat RNA by wheat enzymes, in vitro. For a large fraction of the supernumerary methyl groups that can be introduced into wheat RNA, in vitro, it was not possible to adduce convincing evidence in support of the view that any appreciable quantity of methyl groups is ever introduced at these same sites, in vivo. The possibility that these supernumerary methyl groups might have transient existence, in vivo, and the potential physiological significance of any such occurrence are dealt with as part of a more general discussion of the experimental findings.

Guanosine↗
PubMed1975

Selective labelling ot the methyl carboxylate substituents found in the anticodon sequences of some species of yeast transfer RNA.

(1) By incubation in 0.1 M NaOH for 10 min at room temperature, it is possible to "saponify" some of the methyl carboxylate linkages in bulk yeast tRNA. By incubation with S-adenosyl(Me-14-C)methionine and either homologous (yeast) or heterologous (wheat-embryo) enzymes, it is then possible to "re-esterify" the "saponified" tRNA and thereby effect selective labelling at 5-carboxymethyluridine (Me-14-C)methyl ester residues. (2) There is also selective labelling at 2-thio-5-carboxymethyluridine (Me-14-C)methyl ester residues when "saponified" yeast tRNA is incubated with S-adenosyl(Me-14-C)methionine and homologous (but not heterologous) enzymes. (3) When selectively labelled yeast tRNA is hydrolyzed by RNase T-1, both 5-carboxymethyluridine (Me-14-C)methyl ester and its 2-thio-analogue are released as part of large oligonucleotides, each of which contains roughly 10 nucleotide residues. (4) There are at least three, and possibly four (Me-14-C)methyl ester-containing oligonucleotides released by RNase T-1 digestion of selectively labelled "saponified" yeast tRNA. A comparison of the chromatographic properties of the different (Me-14-C)oligonucleotides suggests that the same 5-carboxymethyluridine residues are probably targets for both homologous and heterologous enzymes. (5) The properties of the selectively labelled oligonucleotides are consistent with the view that some of them probably are derived from yeast tRNA-3-Glu, tRNA-2-Lys, and tRNA-3-Arg, all of which are known to contain 5-carboxymethyl methyl esters as part of their anticodon sequences.

Alkaline Phosphatase↗
PubMed1975

The probable 'capping' of wheat leaf messenger ribonucleates by 7-methylguanosine.

Experiments have been designed to test the possibility that, as with messenger RNA from animal cells, 7-methylguanosine may be bonded through a pyrophosphate bridge to the 5'-termini of messenger RNA molecules from the cells of a higher-plant organism. It is concluded that 7-methylguanosine is probably bonded through a pyrophosphate bridge to the 5'-termini of messenger RNA molecules from wheat leaves.

Binding Sites↗