Rat lens beta-crystallins are internally duplicated and homologous to gamma-crystallins

The nucleotide sequence of two cloned rat lens beta-crystallin cDNAs pRL beta B3-2 and pRL beta B1-3 has been determined. pRL beta B3-2 contains the complete coding information for a beta-crystallin, designated beta B3, of 210 amino acid residues. pRL beta B1-3 is incomplete at its 5' end; the 5' codogenic information which is not present in this cDNA clone was deduced from the cloned gene. pRL beta B1-3 codes for a beta-crystallin polypeptide, designated beta B1, whose full length is 247 amino acid residues. Considerable sequence homology is noted between the amino- and carboxy-terminal halves of each protein. The two rat beta-crystallins show a substantial sequence homology with each other (60%) as well as with the published sequences of rat gamma-crystallin (37%) and bovine and murine beta-crystallins (55 and 45%). All these proteins have a two-domain structure which, like the bovine gamma II-crystallin, might be folded into four remarkably similar protein motifs. Our data further indicate that the beta-crystallins can be subdivided into two groups which are evolutionarily related. Both groups are, although more distantly, also related to the gamma-crystallins.     

Carbamoyl-phosphate synthetases from Neurospora crassa. Immunological relatedness of the enzymes from Neurospora, bacteria, yeast, and mammals

Neurospora crassa contains two carbamoyl-phosphate synthetases: a mitochondrial enzyme (CPS-A) which supplies carbamoyl phosphate for arginine biosynthesis, and a nuclear enzyme whose product is used for the synthesis of pyrimidines. We have prepared antiserum against a highly purified preparation of the large subunit of CPS-A and have used the antiserum to demonstrate that the large subunit is, like most mitochondrially localized proteins, initially synthesized as a higher molecular weight precursor. The CPS-A antiserum cross-reacts with the nuclear enzyme, allowing us to identify the product of the complex N. crassa pyr-3 genetic locus as a protein with a subunit molecular weight of 180,000. Finally, we have found that the CPS-A antiserum also cross-reacts with carbamoyl-phosphate synthetases from bacteria, yeast, and mammals. The immunological relatedness of carbamoyl-phosphate synthetases from such diverse species suggests that the protein sequences required for carbamoyl phosphate production have been highly conserved during the course of evolution.     

Role of carnitine during development

Fatty acids are an important fuel source for neonates. The utilization of long chain fatty acids as a fuel source is dependent upon adequate concentrations of carnitine. Carnitine also has functions in other physiological processes critical to the survival of the neonate such as lipolysis, thermogenesis, ketogenesis, and possibly regulation of certain aspects of nitrogen metabolism. Plasma and tissue carnitine concentrations in neonates are depressed compared with those of older individuals. The capability for carnitine biosynthesis is much less in the neonate than in the adult. Human milk contains carnitine and appears to be the major source of carnitine to meet the neonate's metabolic needs. However, total parenteral nutrition solutions and soy-based infant formulas contain no carnitine. Evidence is accumulating that all infant diets may need to supply carnitine to meet the neonate's metabolic needs.     

Natural relationship between bacteroides and flavobacteria.

Comparisons among 16S rRNA sequences from various eubacteria reveal a natural relationship between the bacteroides (represented by the Bacteroides fragilis sequence) and a phylogenetic unit that comprises the flavobacteria, cytophagae, flexibacteria, and others (represented by the Flavobacterium heparinum sequence). Although the relationship is not a close one, it is, nevertheless, specific. rRNAs from these two organisms are not only closer to one another in overall sequence than they are to outgroup species (such as Bacillus subtilis, Escherichia coli, Desulfovibrio desulfuricans, and Agrobacterium tumefaciens), but they show common idiosyncrasies (i.e., derived characteristics) in both rRNA sequences and higher-order structures.     

A new method for rapid assignment of S-S bridges in proteins

A new method for complementing existing protein chemical techniques for the assignment of S-S bridge positions in amino-acid sequences is described. The principle of the method is the direct examination of the masses of protein fragments, obtained by chemical or enzymatic degradation. Proteins are digested under conditions known to minimise disulphide reduction and reshuffling, and the unfractionated digest is examined directly by high field magnet (or other high mass) fast atom bombardment or Californium mass spectrometry. Disulphide linked peptides are identified from their unique masses, and by comparison with the spectrum of digested and reduced samples in which the signal corresponding to the S-S linked peptide(s) is replaced by two signals corresponding to the respective thiol peptide components, if INTER-bridged, or shifted by two mass units (dithiol) if INTRA-bridged. This rapid procedure has considerable potential in assisting with studies on the primary structure of proteins, in crystallographic studies and the monitoring of denaturation/renaturation of recombinant proteins.     

Chemical modification of the Na+/H+ exchanger of thymic lymphocytes. Inhibition by N-ethylmaleimide

A Na+/H+ exchanger is involved in the regulation of cytoplasmic pH and cellular volume in a variety of cells. Little is known about the molecular nature of this exchanger. The purpose of this study was to survey a variety of group-specific covalent reagents as potential inhibitors of the exchanger. Na+/H+ countertransport activity was assayed as the amiloride-sensitive rate of Na+-induced alkalinization in acid-loaded lymphocytes, or as the rate of swelling in cells suspended in sodium propionate medium. Activity was not affected by proteinases or by carboxyl-group and amino-group specific reagents. A significant inhibition was produced by diethylpyrocarbonate, a histidine-specific reagent and by N-ethylmaleimide, a sulfhydryl group reagent. A similarly reactive but nonpermeating sulfhydryl agent, glutathione-maleimide, failed to inhibit Na+-H+ exchange. Moreover, the reaction with N-ethylmaleimide was sensitive to changes in the cytoplasmic pH. The data suggest that the chemically reactive groups of the Na+/H+ exchanger of lymphocytes have limited exposure to the extracellular medium but that an internally located sulfhydryl group is critical for the cation-exchange activity.     

Cultivation of Neisseria meningitidis serogroups A, B and C in a synthetic nutrient broth

The processes of the cultivation of N. meningitidis, serogroups A, B and C, in a liquid synthetic culture medium have been studied. Strictly group-specific biomass has been obtained. The maximum productivity at all stages of the batch cultivation of N. meningitidis strains 125 and 133 in this medium does not differ from that at similar stages of cultivation in modified Cohen-Wheller semisynthetic medium. In the serotype antigen preparations obtained from N. meningitidis strain 125 grown in the above-mentioned liquid synthetic culture medium basic polypeptides with a molecular weight of 33000, 36000 and 41000 D have been detected. Their presence in N. meningitidis cells is linked with the growth phase of the population.     

Effect of cortisol, its polymeric derivative and adenylate cyclase activators on the content of cyclic AMP in rat thymocytes

In has been shown that cortisol immobilized on polyvinylpyrrolidone (PVP-GC) affects cyclic AMP production stimulated by adenosine and isoproterenol in rat thymocytes. This effect of PVP-GC is specific for cortisol: antiglucocorticoid progesterone (at a concentration of 10(-5) M) inhibited completely the action of PVP-GC on the intracellular cAMP level. It is suggested that cortisol effect on cAMP production is one of the mechanisms of glucocorticoid hormone action in target cells.