Identification, purification, and characterization of phosphotyrosine-specific protein phosphatases from cultured chicken embryo fibroblasts.

Tyrosine phosphorylation catalyzed by a unique class of protein kinases is an important process in both normal cell proliferation and oncogenic transformation. In this study, phosphoprotein phosphatases specific for the dephosphorylation of phosphotyrosine residues were partially purified from secondary chicken embryo fibroblasts, using 32P-labeled immunoglobulin G phosphorylated by pp60src as substrate. Crude cell extracts contained ca. 70% of the activity in the soluble form and ca. 30% associated with a crude membrane fraction. The soluble activity was purified by using DEAE-cellulose and carboxymethyl cellulose column chromatography and gel filtration, and at least three enzyme species of apparent Mr 55,000 (pTPI), 50,000 (pTPII), and 95,000 (pTPIII)--comprising ca. 20, 45, and 35%, respectively, of the total activity--were resolved. All three enzymes possessed somewhat similar properties. They had a pH optimum of about 7.4, they were inhibited by Zn2+, vanadate, ATP, and ADP, and they were unaffected by divalent metal cations, EDTA, and F- under standard assay conditions employing a physiological ionic strength. These properties suggest that they represent a class of enzymes distinct from well-known phosphoseryl-phosphothreonyl-protein phosphatases and that dephosphorylation of phosphotyrosine-containing proteins may be carried out by a unique family of phosphoprotein phosphatases. Transformation by Rous sarcoma virus resulted in a small increase in phosphotyrosyl-protein phosphatase activity.     

An inducible pathway is required for mutagenesis in Salmonella typhimurium LT2.

UV mutability of Salmonella typhimurium LT2 was eliminated in the presence of a multicopy plasmid carrying the Escherichia coli lexA+ gene. This result suggests that inducible, SOS-like functions are required for UV mutagenesis in S. typhimurium. S. typhimurium strains carrying either point or deletion mutations in topA had previously been shown to lose their mutability by UV or methyl methanesulfonate (K. Overbye and P. Margolin, J. Bacteriol. 146:170-178, 1981; K. Overbye, S. M. Basu, and P. Margolin, Cold Spring Harbor Symp. Quant. Biol. 47:785-791, 1983). Mitomycin C induction of the phi(mucB'-lacZ') fusion (a DNA damage-inducible locus carried on plasmid pSE205) in S. typhimurium topA was normal, suggesting that RecA is activated in topA mutants. These observations lead us to deduce that S. typhimurium has at least one DNA damage-inducible locus in addition to recA that is required for UV mutability.     

Linear mapping of tryptophan residues in Vesiculovirus M and N proteins by partial chemical cleavage.

Nonlimit chemical cleavage at tryptophan residues of protein labeled at the amino terminus afforded a simple procedure for generating specific fragments and for mapping tryptophan positions. A comparison of the matrix (M) and nucleocapsid (N) proteins of four members of the Vesiculovirus group by this procedure suggests considerable conservation of tryptophan number and location in the four serotypes examined.     

Functional significance of manganese catalase in Lactobacillus plantarum.

A strain of Lactobacillus plantarum which was unable to produce manganese (Mn)catalase (ATCC 8014) grew somewhat more rapidly and to a slightly higher plateau density than did an Mn catalase-positive strain (ATCC 14421), and this was the case during aerobic or anaerobic growth. However, when maintenance of viability was measured during the stationary phase of the growth cycle, the advantage provided by Mn catalase was obvious. Thus, the viability of ATCC 14431 was undiminished over 21 h of aerobic incubation, during the stationary phase, whereas that of ATCC 8014 decreased by seven orders of magnitude. Addition of catalase to the medium or growth in the presence of hemin, which allows catalase synthesis, protected ATCC 8014 against this loss of viability. Suppression of Mn catalase within ATCC 14431 by treatment with NH2OH caused the cells to lose viability when exposed to 4 mM H2O2.     

Genetic evidence that the putative receptor binding domain of apolipoprotein B (residues 3130 to 3630) is not the only region of the protein involved in interaction with the low density lipoprotein receptor

We have searched for sequence differences in the region of the apolipoprotein B (apo B) gene encoding amino acids 3130-3630 in eight individuals with reduced affinity of low density lipoprotein (LDL) for the normal LDL-receptor. All individuals were hypercholesterolaemic and were selected either on the basis of reduced fractional catabolic rate (FCR) of autologous LDL or substantially reduced binding of their LDL to normal LDL-receptors determined by an in vitro cell growth assay using the U937 macrophage-like cell line. Segments of the apo B gene were amplified by the polymerase chain reaction. Using a combination of cloning and sequencing the amplified fragment, together with chemical cleavage mismatch analysis, no sequence differences were identified in this region of the gene. We therefore conclude that variation outside the region of the apo B gene that codes for amino acids 3130-3630 must be responsible for the reduced LDL clearance in these patients.     

Few intra-lake variations of physiological parameters in perch, Perca fluviatilis

Intra-lake variations in physiological parameters, representing haematology, plasma ion composition and carbohydrate metabolism, were investigated in perch (Perca fluviatilis), inhabiting a comparatively unpolluted lake. Provided the perch were subjected to a standardized procedure for capture, handling, recovery after capture, and sampling, only few and minor differences were observed in 21 parameters investigated when 3 groups were compared to a control group of perch. It is concluded that the experimental design used is suitable for the examination of the physiological status of perch in the field.