Molecular mapping of murine I region recombinants. III. Crossing over at two discrete sites within the beta 1-beta 2 intron of the E beta gene

The murine MHC provides a unique genetic system for studying meiotic recombination. A large number of murine H-2 recombinants cross over within a stretch of the E beta gene referred to as the E beta hot spot. The crossing over of eight such recombinants, derived from the s and k haplotypes, was studied at the nucleotide level. A 3-kb stretch of DNA, 3' to the beta 1 exon of the E beta gene, was sequenced after amplification of the genomic DNA from B10.S (one of the parental strains) by polymerase chain reaction. A number of sequence variations were identified with respect to B10.A (the other parental strain). Examination of these sequence variations by RFLP, simple sequence length polymorphism, as well as direct sequencing after polymerase chain reaction-amplification of genomic DNA from the recombinants led to unambiguous identification of the cross-over sites. Although all eight recombinants crossed over within the beta 1-beta 2 intron, two discrete nonoverlapping sites were involved. Five of the recombinants B10.BASR1, B10.ASR1, B10.ASR12, B10.HTT, and B10.S(9R) crossed over within a maximum of 395 bp of DNA 3' to the beta 1 exon. The remaining three recombinants B10.ASR7, B10.ASR11, and B10.S(8R) crossed over within 950 bp of DNA, adjacent to the cross-over site noted above. Each of these stretches of DNA was completely identical in the two parental haplotypes precluding further dissection of the cross-over sites. These cross-over sites are within those reported for the b and k recombination.     

Apolipoprotein H (beta-2-glycoprotein I) polymorphism in Asians.

Apolipoprotein H (APOH) (beta-2-glycoprotein I) polymorphism has been studied in 1159 Asians. The sample included 872 Chinese, 179 Asiatic Indians (Dravidian), 91 Filipinos, and 17 Malays. APOH polymorphism was determined by isoelectric focusing of sera in thin-layer polyacrylamide gels containing 3 M urea followed by immunoblotting. The frequencies of the three alleles--APOH*1, APOH*2, and APOH*3--were found to be 0.031, 0.900, and 0.069 in the Chinese; 0.061, 0.866, and 0.073 in the Dravidian Indians; 0.055, 0.923, and 0.022 in the Filipinos; and 0.088, 0.882, and 0.029 in the Malays. The phenotypic distribution was at Hardy-Weinberg equilibrium in all the populations.     

Radiation-induced inactivation of dihydroorotate dehydrogenase in dilute aqueous solution.

The inactivation of dihydroorotate dehydrogenase by gamma irradiation in dilute aqueous solution has been investigated. The activity of the enzyme decreased exponentially as a function of the absorbed dose under aerated and nitrous oxide-saturated conditions. The contributions of the individual radical species derived from water radiolysis were estimated from the inactivation results observed under aerated, argon-saturated, and nitrous oxide-saturated conditions. The hydrogen atom and hydroxyl radical were found to be important in enzyme inactivation. The effect of selected inorganic radical anions such as Br.2-, I.2-, and (SCN).2- on the enzyme activity was also studied, and the results implicate the possible involvement of cysteine and tyrosine residues in the catalytic activity of dihydroorotate dehydrogenase. Changes in the kinetic parameters (Michaelis-Menten constant, Km, and maximal velocity, Vmax) due to irradiation under the conditions investigated suggest that radiation-induced inactivation is due to modification of the substrate binding sites and that of the active site residues in the enzyme. Evidence for the reduction of iron-sulfur centers in the enzyme during the inactivation process has been put forward from the difference spectrum of the irradiated dihydroorotate dehydrogenase. It has also been shown by electrophoretic studies that radiation-induced inactivation was not due to any fragmentation of the protein structure or the formation of any intermolecular crosslinking.     

Apolipoprotein A-IV polymorphism in Singapore ethnic groups

A total of 627 subjects comprising 455 Chinese, 127 Dravidian Indians and 45 Malays were investigated for serum Apo A-IV polymorphism. The frequency of Apo A-IV*2 was found to be significantly higher (p less than 0.001) in Indians (0.043) compared to that in the Chinese (0.010) and Malays (0.011). The frequency of A-IV*3 was found to be around 0.02 in all the ethnic groups. A low frequency of A-IV*4 (less than 0.01) was observed in the Chinese and Indians. The phenotypic distribution of Apo A-IV was at Hardy-Weinberg equilibrium in the three ethnic groups.     

Studies on organ weights in naproxen treated rats after intermittent exposure to simulated high altitude

Rats were exposed intermittently for 8h per day over 6 days at simulated high altitude of 20 000 feet. One group of altitude-exposed animals was treated with naproxen, a prostaglandin inhibiting drug. Significant reduction in body weight gain was observed in both altitude-exposed and drug-treated altitude-exposed animals compared to the control group. Right and left ventricular weights and weights of the adrenal glands were increased significantly in altitude-exposed and altitude-exposed drug-treated animals. The weight of the spleen was increased significantly in altitude-exposed animals whereas no such increase of splenic weight was observed in drug-treated altitude-exposed group of animals. On the other hand, the weight of the liver was decreased significantly in both cases. In drug-treated altitude-exposed animals, the unaltered splenic weight was thought to be due to inhibition of the erythropoietic activity.     

Substrate competition and specificity at the active site of amylopullulanase from Clostridium thermohydrosulfuricum

A highly thermostable pullulanase purified from Clostridium thermohydrosulfuricum strain 39E displayed dual activity with respect to glycosidic bond cleavage. The enzyme cleaved alpha-1,6 bonds in pullulan, while it showed alpha-1,4 activity against malto-oligosaccharides. Kinetic analysis of the purified enzyme in a system which contained both pullulan and amylose as the two competing substrates were used to distinguish the dual specificity of the enzyme from the single substrate specificity known for pullulanases and alpha-amylases.     

Cloning and expression of the Clostridium thermosulfurogenes glucose isomerase gene in Escherichia coli and Bacillus subtilis.

The gene that encodes thermostable glucose isomerase in Clostridium thermosulfurogenes was cloned by complementation of glucose isomerase activity in a xylA mutant of Escherichia coli. A new assay method for thermostable glucose isomerase activity on agar plates, using a top agar mixture containing fructose, glucose oxidase, peroxidase, and benzidine, was developed. One positive clone, carrying plasmid pCGI38, was isolated from a cosmid library of C. thermosulfurogenes DNA. The plasmid was further subcloned into a Bacillus cloning vector, pTB523, to generate shuttle plasmid pMLG1, which is able to replicate in both E. coli and Bacillus subtilis. Expression of the thermostable glucose isomerase gene in both species was constitutive, whereas synthesis of the enzyme in C. thermosulfurogenes was inducible by D-xylose. B. subtilis and E. coli produced higher levels of thermostable glucose isomerase (1.54 and 0.46 U/mg of protein, respectively) than did C. thermosulfurogenes (0.29 U/mg of protein). The glucose isomerases synthesized in E. coli and B. subtilis were purified to homogeneity and displayed properties (subunit Mr, 50,000; tetrameric molecular structure; thermostability; metal ion requirement; and apparent temperature and pH optima) identical to those of the native enzyme purified from C. thermosulfurogenes. Simple heat treatment of crude extracts from E. coli and B. subtilis cells carrying the recombinant plasmid at 85 degrees C for 15 min generated 80% pure glucose isomerase. The maximum conversion yield of glucose (35%, wt/wt) to fructose with the thermostable glucose isomerase (10.8 U/g of dry substrate) was 52% at pH 7.0 and 70 degrees C.     

Induction of carboxymethylcellulase inMacrophomina phaseolina

Macrophomina phaseolina, the well-known jute pathogenic fungus produces very low levels of both extra- and intracellular carboxymethylcellulase even in the absence of any cellulose as carbon source in the medium. However, the production of these enzymes is greatly induced by soluble carboxymethylcellulose. The carboxymethylcellulase inM.phaseolina is repressed by glucose.     

Bimodal oxygen uptake in juveniles and adults amphibious fish,Channa (= Ophiocephalus) marulius

Oxygen uptake of Channa marulius was studied under water with and without access to air. There was a significant increase in the oxygen uptake through the gills when access to air was prevented. However, this value (0.863 ± 0.058 mlO₂/indiv./h) was quite low in comparison to the total bimodal oxygen uptake (2.04 ± 0.14 mlO₂/indiv./h) in juveniles. In adult fish the oxygen uptake per unit time increased appreciably (4.673 ± 0.404 mlO₂/indiv./h). In juveniles as well as in adults the air breathing dominated over aquatic breathing. This fish showed a definite circadian rhythm in the bimodal oxygen uptake during different hours of the day.This work was performed in the Ichthyology Laboratory, P. G. Dept. of Zoology, Bhagalpur University, and was supported by a research grant from Bhagalpur University