Bacteriophage Ø105clz induces the GroEL-homologue protein inBacillus subtilis

Using two-dimensional polyacrylamide gel electrophoresis, the GroEL homologue ofBacillus subtilis was shown to be induced upon infection with Ø105clz, a clear plaque mutant of the temperate bacteriophage Ø105. Western blotting of one dimensional polyacrylamide gels also showed the induction of the GroEL homologue when cells were infected with Ø105clz.     

THE INFLUENCE OF THE MATING SYSTEM ON SEED SIZE VARIATION IN CRINUM ERUBESCENS (AMARYLLIDACEAE)

We present evidence that extreme seed size variation in fruits of Crinum erubescens (range: 0.1 to 66.5 g per seed) occurs when mating pairs are inbred, either from selfing or biparental inbreeding. Several relatively uniform seeds of intermediate size are produced when pollen from several pollen donors is applied simultaneously to a flower. Selfed fruits and some fruits pollinated with a single pollen donor produce both large and small seeds, although selfed fruits produce fewer seeds than outcrossed fruit. These results are contrary to the hypothesis that variation in seed size is attributable to either pollen competition or differential allocation of maternal resource to seeds of different genotypes.     

cDNA and protein sequence of a major pea seed albumin (PA 2 : Mr≈26 000)

Summary Pea albumin 2 (PA2:M_r26000) is a major component of the albumin fraction derived from aqueous salt extracts of pea seed. Sodium dodecylsulfate-polyacrylamide gel electrophoresis and chromatography on DEAE-Sephacel resolve PA2 into two closely related components (PA2a and PA2b). A cDNA clone coding for one of these components has been sequenced and the deduced amino acid sequence compared with partial, chemically-determined sequences for cyanogen bromide peptides from both PA2 components. Complete amino acid sequences were obtained for the C-terminal peptides. The PA2 molecule of 230 amino acids contains four imperfect repeat sequences each of approximately 57 amino acids in length.The combined sequence data, together with a comparison of PA2-related polypeptides produced in vitro and in vivo, indicate that PA2 is synthesized without a signal sequence and does not undergo significant post-translational modification. Although both forms of PA2 contain Asn-X-Thr consensus sequences, neither form is glycosylated. Accumulation of PA2 contributes approximately 11% of the sulfur-amino acids in pea seeds (cysteine plus methionine equals 2.6 residues percent). Suppression of levels of PA2 polypeptides and their mRNAs in developing seeds of sulfur-deficient plants is less marked than that for legumin, in spite of the lower content of sulfur-amino acids in legumin.     

Construction and properties of a chimeric bacteriophage lysis gene

Abstract The genomes of DNA phage ΦX174 and of RNA phage MS2 each encode a single lysis protein, E protein and L protein, respectively. Based on the known DNA and protein sequences, and with the aid of structural predictions of the proteins, a chimeric gene was constructed. The resulting protein was composed of the N-terminal sequence of E protein and the C-terminal sequence of L protein. The truncated E and L polypeptides used in this construction were functionally inactive. However, the chimeric gene product had very high lysis-inducing activity. This construction is an example which could be extended to the engineering of other lysis proteins designed with specific biotechnological processes in mind.     

Aminoglycoside suppression at UAG, UAA and UGA codons in Escherichia coli and human tissue culture cells

Summary We have compared the suppression of nonsense mutations by aminoglycoside antibiotics inEscherichia coli and in human 293 cells. Six nonsense alleles of the chloramphenicol acetyl transferase (cat) gene, in the vector pRSVcat, were suppressed by growth in G418 and paromomycin. Readthrough at UAG, UAA and UGA codons was monitored with enzyme assays for chloramphenicol acetyl transferase (CAT), in stably transformed bacteria and during transient expression from the same plasmid in human 293 tissue culture cells. We have found significant differences in the degree of suppression amongst three UAG codons and two UAA codons in different mRNA contexts. However, the pattern of these effects are not the same in the two organisms. Our data suggest that context effects of nonsense suppression may operate under different rules inE. coli and human cells.     

Critical evaluation of thein situ nitrate reductase assay

Anin situ method, derived from anin vivo method, was used to determine nitrate reductase activity (NRA) in:i) excised barley and corn shoots and excised soybean leaves during a N-depletion experiment and; ii) roots and shoots of N-depleted barley and corn seedlings during induction of nitrate, reductase (NR). Nitrate reduction, calculated from thesein situ RNA measurements, was compared with estimates of each organ's nitrate reduction in light aerobic conditions from NO ₃ ⁻ consumption and a¹⁵N model (Gojonet al., 1986b). Thein situ RNA of roots strongly underestimated their¹⁵NO ₃ ⁻ reduction. In contrast, in barley and corn shoots and in the first trifoliolate leaves from 26-day-old, soybean, thein situ NRA assay gave a fair approximation of the true NO ₃ ⁻ reduction rate (relative differences ranging from −14 to +32%). In young soybean leaves (from 20-day-old plants), however, thein situ NRA strongly underestimated the actual NO ₃ ⁻ reduction. The physiological significance of thein situ NRA assay in shoots and roots, and its value for field studies are discussed from these results.     

Consequences of terbium (111) binding on the conformation and enzymatic activity of Guinea pig liver transglutaminase

Calcium ions are crucial for expression of transglutaminase activity. Although lanthanides have been reported to substitute for calcium in a variety of protein functions, they did not replace the calcium requirement during transglutaminase activity measurements. Furthermore, lanthanides strongly inhibited purified liver transglutaminase activity using either casein or fibrinogen as substrates. Terbium (III) inhibition of transglutaminase-catalyzed putrescine incorporation into casein was not reversed by the presence of 10–200 fold molar excess of calcium ions (Ki for Tb(III)=60 µM). Conformational changes in purified liver transglutaminase upon Tb(III) binding were evident from a biphasic effect of Tb(III) on transglutaminase binding to fibrin. Low concentrations of Tb(III) (1 µM to 10 µM inhibited the binding of transglutaminase to fibrin, whereas higher concentrations (20 µM to 100 µM promoted binding. Conformational changes in purified liver transglutaminase consequent to Tb(III) binding were also demonstrated by fluorescence spectroscopy due to Forster energy transfer. Fluorescence emission was stable to the presence of 200 mM NaCl and 100 mM CaCl₂ only partially quenched emission. Purified liver transglutaminase strongly bound to Tb(III)-Chelating Sepharose beads and binding could not be disrupted by 100 mM CaCl₂ solution. Our data suggest that Tb(III)-induced conformational changes in transglutaminase are responsible for the observed effects on enzyme structure and function. The potential applications of Tb(III)-transglutaminase interactions in elucidating the structure-function relationships of liver transglutaminase are discussed.     

The effect of glucose-6-phosphate isomerase genotype onin vitro specific activity andin vivo flux inMytilus edulis

Four samples of the musselMytilus edulis were taken between 1984 and 1987 from Stony Brook, New York, and used to study the glucose-6-phosphate isomerase (GPI) polymorphism in this species.In vitro specific activity andin vivo flux measured in the same animals were found to be significantly correlated. A significant effect of GPI genotype on flux was observed in one of the samples; overall, significant evidence of effect of genotype on enzyme activity was also obtained. GPI activities of common genotypes tend to deviate less from the population mean than those of rare (frequency less than 5%) genotypes. This suggests the possibility that rare GPI genotypes are rare as a consequence of having biochemical properties that deviate from an optimum level and, therefore, having a lower fitness. In support of this hypothesis, we found in one of our samples that shell length is a concave function of GPI activity with an intermediate optimum activity level. The financial support provided to P.J.N.S. by the Luso-American Educational Commission (Fulbright Program), the Instituto Nacional de Investigacao Científica (Portugal), and the Faculdade de Ciências da Universidade de Lisboa during several stages of this research is gratefully acknowledged. Financial support from the Ministerio de Educatión y Ciencia (Spain) in the form of a postdoctoral Fulbright/MEC fellowship to M.S. is also gratefully acknowledged. Research was supported by National Science Foundation Grant BSR-8415060 to R.K.K. This is contribution No. 736 from the Program in Ecology and Evolution, State University of New York at Stony Brook. On leave from Departamento de Biologia Vegetal, Faculdade de Ciências, Universidade de Lisboa, Campo Grande C2, Lisboa, Portugal.