In vitro effects of gossypol on testicular lactic dehydrogenase-X and other dehydrogenases

Thein vitro inhibition of several rat testis dehydrogenases by gossyPol was examined. Inclusion of the coenzyme (substrate for NADP⁺-isocitrate dehydrogenase) in the Preincubation mixture containing the enzyme and gossyPol, Protected the enzymes against inhibition by gossyPol. Lactic dehydrogenase-X was amongst the least Protected enzymes. This, couPled with its lowK _i for gossyPol makes it one of the most vulnerable target enzymesin vivo for gossyPol action. The inhibition kinetics for lactic dehydrogenase-X were comPetitive when NADH was Present during Preincubation, but non-comPetitive when the coenzyme was excluded during Preincubation. In the latter condition, the enzyme seems to undergo Progressive inactivation with time causing a nonreversible tyPe of inhibition.     

Raine syndrome

Raine syndrome is a rare genetic disorder with characteristic features of exophthalmos, choanal atresia or stenosis, osteosclerosis and cerebral calcifications. Most of babies with this disorder die immediately after birth. We report a baby who was 7 weeks old at the time of presentation.     

A Versatile Microassay for Elastase Using Succinylated Elastin

We have developed a rapid, versatile, and sensitive elastase assay that is based on the measurement of primary amines that are exposed due to enzymatic degradation of proteins, using succinylated elastin as the substrate for elastase. After incubation with elastase the degree of digestion is determined with trinitrobenzene sulfonic acid. The assay is rapid and sensitive, detecting elastase down to 1 ng/ml, and is linear up to enzyme concentrations of 10 μg/ml. The assay is carried out in microtiter plate wells and therefore offers the potential for assaying numerous samples of small volume. The use of succinylated elastin shows specificity for elastase over the control protease, trypsin. This assay is also versatile because it can be applied to samples such as cell culture supernatants, blood plasma, tissue biopsies, and tissue homogenates.     

Intramolecular cross-linking of insulin. Preparation and properties of oxalyl- and malonyl-bis(methionyl) insulin

The bifunctional reagents, oxalyl-(Met-ONp)2 and malonyl-(Met-ONp)2 have been prepared and investigated as reversible cross-linking reagents for insulin and model compounds. The removal of the cross-linking residues was demonstrated by the cyanogen bromide cleavage of oxalyl-(Met-Phe-OMe)2 and malonyl-(Met-Phe-OMe)2. Zinc-insulin reacted with a molar equivalent of oxalyl-(Met-ONp)2 or malonyl-(Met-ONp)2 in presence of excess triethylamine to yield oxalyl-(Met)2-insulin and malonyl-(Met)2-insulin, respectively. In these derivatives the N-terminal phenylalanine (B1 residue) was free. Thus the cross-link was between A1 and B29 residues in insulin. All three disulfide bonds of these insulin derivatives undergo reduction with tributylphosphine to give six sulfhydryls. Air-oxidation of reduced oxalyl-(Met)2-insulin and malonyl-(Met)2-insulin in 0.05 M disodium phosphate, pH 9.5, yielded products which were indistinguishable from oxalyl-(Met)2-insulin and malonyl-(Met)2-insulin respectively, as measured by physicochemical and biological methods. Cyanogen bromide cleavage of reduced and reoxidized malonyl-(Met)2-insulin in 70% formic acid regenerated insulin quantitatively, but only 40% of insulin was determined from similar treatment of oxalyl-(Met)2-insulin. The regenerated insulins exhibited the biological activity of native insulin. These studies strongly suggest that disulfide bonds formed during oxidation of reduced oxalyl-(Met)2-insulin and malonyl-(Met)2-insulin are identical to those found in insulin.     

Cloning and expression of MPT83 gene from Mycobacterium tuberculosis in E. coli BL21 as vaccine candidate of tuberculosis: A preliminary study

The appearance of Mycobacterium tuberculosis strains leading to drug resistance has caused new problems in TB treatment in various parts of the world and forces WHO to declare TB as a global emergency. With the increase of TB drug resistance, it is convinced that a more effective vaccine development will stop the epidemic of TB. Some M. tuberculosis antigens, one of which is MPT83, have been examined as TB vaccine candidate. MPT83 antigen, which is very immunogenic in lipoprotein micro bacteria, is identified as surface cell interrelated to antigen with cytometry circulation. Having TB resistance from BCG vaccine, MPT83 is considered TB vaccine candidate that can protect people against TB at adult age. The purpose of this research is to conduct amplification of MPT83 antigen cloning, and expression of its antigen on E. coli bacteria. From the result of the research, it is expected that raw material to produce TB vaccine as well as a high-quality antigen can be obtained. The band of DNA in PCR product is 660?bp, while the one in pGEMT-Easy-Mpt83 recombinant plasmid is 3678?bp. This is expressed in E. coli BL21 strain and produces 48?kDa protein as well as GST-MPT83 fusion protein.