Protein Synthesis in Mycobacterium tuberculosis H37Rv and the Effect of Streptomycin in Streptomycin-Susceptible and -Resistant Strains

An efficient in vitro amino acid-incorporating system from Mycobacterium tuberculosis H37Rv was standardized. Ribonucleic acid (RNA) isolated from phage-infected M. smegmatis cells served as natural messenger RNA and directed the incorporation of 14C-amino acids into protein. The effects of various antitubercular drugs and “known inhibitors” of protein synthesis on amino acid incorporation were studied. Antibiotics like chloramphenicol and tetracycline inhibited mycobacterial protein synthesis, though they failed to prevent the growth of the organism. This failure was shown to be due to the impermeability of mycobacteria to these drugs by use of “membrane-active” agents along with the antibiotics in growth inhibition studies. Several independent streptomycin-resistant mutants of M. tuberculosis H37Rv were isolated. Streptomycin inhibited the incorporation of 14C-amino acids into proteins by whole cells of a streptomycin-susceptible strain by more than 90%, whereas very little or no inhibition was observed in either high-level or low-level streptomycin-resistant strains.

Purification, crystallization and partial characterization of the antitumour and insecticidal protein subunit from the delta-endotoxin of Bacillus thuringiensis var. thuringiensis

The antitumour protein from the α-endotoxin of Bacillus thuringiensis var. thuringiensis has been purified, crystallized and partially characterized. The same protein also shows the insecticidal activity. According to amino acid analysis it is an acidic protein with a molecular weight of approx. 13 000.

Purification, crystallization and partial characterization of the antitumour and insecticidal protein subunit from the σ-endotoxin of Bacillus thuringiensis var. thuringiensisstar, open

The antitumour protein from the α-endotoxin of Bacillus thuringiensis var. thuringiensis has been purified, crystallized and partially characterized. The same protein also shows the insecticidal activity. According to amino acid analysis it is an acidic protein with a molecular weight of approx. 13 000.

Pregnancy suppression by active immunization against gestation-specific riboflavin carrier protein

A riboflavin carrier protein isolated from chickens cross-reacts with a gestation-specific rodent carrier for riboflavin. Active immunization of female rats of proved fertility with the purified chicken carrier protein completely yet reversibly suppressed early pregnancy without impairing implantation per se. Concurrently there were no discernible adverse effects on maternal health in terms of weight gain, vitamin status, and fertility.

Protein A: nature's universal anti-antibody

Staphylococcal protein A specifically interacts with immunogobulins. This fact is being used in various disciplines of biology and some of the unique properties of protein A and their applications are summarized in this review.

Nature of the thiamin-binding protein from chicken egg yolk

A simple, rapid and efficient procedure for the purification of thiamin-binding protein from chicken egg yolk was developed. The method involved removal, by exclusion, of lipoproteins from DEAE-cellulose and subsequent elution of water-soluble proteins held on the ion-exchanger with 1 M-NaCl, followed by treatment of the eluted protein fraction with an aqueous suspension of dextran/charcoal to generate apoprotein from the holoprotein. The resultant protein fraction was subjected to bioaffinity chromatography on thiamin pyrophosphate--AE (aminoethyl)-Sepharose. The protein eluted specifically with 10 microM-thiamin at pH 7.0, was homogeneous by the criteria of polyacrylamide-gel disc electrophoresis, had a mol.wt. of 38 000 +/- 2000 and was not a glycoprotein. The purified thiamin-binding protein specifically interacted with riboflavin-binding protein with no detectable deleterious affect on its (14C)thiamin-binding capacity. The protein bound [14C]thiamin with a molar ratio of 1.0, with dissociation constant (Kd) 0.41 microM. This protein-ligand interaction was inhibited by thiamin analogues and antagonists. The absorption spectrum of the protein in the presence of thiamin exhibited significant hypochromism at the 278 nm band, indicating the involvement of aromatic amino acid residues of the protein, during its binding to the ligand. The protein cross-reacted with the monospecific antiserum to egg-white thiamin-binding protein, showing thereby that thiamin-binding proteins present in chicken egg yolk and white are the products of the same structural gene.

A fast method of comparing protein structures

Comparative studies on protein structures form an integral part of protein crystallography. Here, a fast method of comparing protein structures is presented. Protein structures are represented as a set of secondary structural elements. The method also provides information regarding preferred packing arrangements and evolutionary dynamics of secondary structural elements. This information is not easily obtained from previous methods. In contrast to those methods, the present one can be used only for proteins with some secondary structure. The method is illustrated with globin folds, cytochromes and dehydrogenases as examples.

Estrogen-induced synthesis of riboflavin-binding protein in immature chicks kinetics and hormonal specificity

The kinetics of estrogen-induced elevation in the plasma concentration of riboflavin-binding protein, a minor yolk constituent, was investigated in immature male chicks, using a specific and sensitive radioimmunoassay proceudre. Following a single injection of the hormone, the plasma riboflavin-binding protein content was enhanced several-fold at 6 h. reaching peak levels around 48 h and declining thereafter. A two-fold amplication of the response was evident on secondary stimulation with the hormone. A 4-h lag phase prior to onset of induction was noticed during both primary and secondary stimulat ions with the steroid hormone. The magnitude of the response was dependent on the hormonal dose whereas the initial lag phase and the time of peak riboflavin-binding protein accumulation were unaltered within the range of hormonal doses tested. The half-life of riboflavin-binding protein in the circulation was 10 h, as calculated from measurement of the rate of disappearance of exogenously administered 125I-labelled protein. Simultaneous administration of progestrone did bot affect the kinetics of riboflavin-binding protein production. On the other hand, the antiestrogens, cis- and trans-clomiphene citrates, given 30 min prior to estrogen and cycloheximide, effectively countered the hormone-induced riboflavin-binding protein elaboration. Both progesterone and the anti-esterogens per se were completely ineffective in substituting for estrogen in the inductive ptrocess.

Mitochondrial localization of functional ferrochelatase from Plasmodium falciparum

In the malarial parasite, enzymes of heme-biosynthetic pathway are distributed in different cellular compartments. The site of localization of ferrochelatase in the malarial parasite is crucial, since it will decide the ultimate site of heme synthesis. Earlier results have differed in terms of localization, being the mitochondrion or apicoplast and the functional enzyme has not been cloned, expressed and characterized. The present study reveals that Plasmodium falciparum ferrochelatase (PfFC) gene encodes multiple transcripts of which the one encoding the full length functional protein (PfFC) has been cloned and the recombinant protein over-expressed and purified from E. coli cells. The enzyme shows maximum activity with iron, while zinc is a poor substrate. Immunofluorescence studies with antibodies to functional ferrochelatase reveal that the native enzyme is localized to the mitochondrion of the parasite indicating that this organelle is the ultimate site of heme synthesis.

Photocytotoxic 3d-Metal Scorpionates with a 1,8-Naphthalimide Chromophore Showing Photoinduced DNA and Protein Cleavage Activity

Ternary 3d-metal complexes of formulation [M(Tp(Ph))(py-nap)](ClO4)(1-3), where M is Co(II) (1), Cu(II) (2), and Zn(II) (3); Tp(Ph) is anionic tris (3-phenylpyrazolyl)borate; and py-nap is a pyridyl ligand with a conjugated 1,8-naphthalimide moiety, have been prepared from the reaction of metal perchlorate with KTp(Ph) and py-nap in CH2Cl2. The complexes have been characterized from analytical and physicochemical data. The complexes are stable in solution as evidenced from the electrospray ionization mass spectrometry data. The complexes show good binding propensity with calf thymus (CT) DNA, giving binding constant (K-b) values of similar to 10(5) M-1 and a molecular ``light-switch'' effect that results in an enhancement of the emission intensity of the naphthalimide chromophore on binding to CT DNA. The complexes do not show any hydrolytic cleavage of DNA. They show poor chemical nuclease activity in the presence of 3-mercaptopropionic acid or hydrogen peroxide (H2O2). The Co(II) and Cu(II) complexes exhibit oxidative pUC19 DNA cleavage activity in UV-A light of 365 rim. The Zn(II) complex shows moderate DNA photocleavage activity at 365 nm. The Cu(II)complex 2 displays photoinduced DNA cleavage activity in red light of 647.1 nm and 676 rim and near-IR light of >750 rim. A mechanistic studyin UV-A and visible light suggests the involvement of the hydroxyl radical as the reactive species in the DNA photocleavage reactions. The complexes also show good bovine serum albumin (BSA) protein binding propensity, giving K-BSA values of similar to 10(5) M-1. Complexes 1 and 2 display significant photoinduced BSA cleavage activity in UV-A light. The Co(II) complex 1 shows a significant photocytotoxic effect in HeLa cervical cancer cells on exposure to UV-A light of 365 nm, giving an IC50 value of 32 mu M. The IC50 value for the py-nap ligand alone is 41.42 mu m in UV-A light. The IC50 value is >200 mu M in the dark, indicating poor dark toxicity of 1. The Cu(II) complex 2 exhibits moderate photocytotoxicity and significant dark toxicity, giving IC50 values of 18.6 mu m and 29.7 mu m in UV-A light and in the dark, respectively.

Oestrogen induction of riboflavin-binding protein in immature chicks. Nature of the secretory protein.

The riboflavin-binding protein isolated from sera of oestrogen-treated male chicks as well as that synthesized and secreted in vitro by the chicken liver have the same molecular size as that of the egg-yolk protein. Functionally the serum and yolk proteins are similar. This is in contrast with the hormone-induced synthesis, secretion and deposition of phosvitin and lipovitellin in the ovary.

Mode of Action of a Purified Antitumor Protein from the Proteinaceous Crystal of Bacillus thuringiensis subsp. thuringiensis on Yoshida Ascites Sarcoma Cells

A purified antitumor protein from the proteinaceous crystal of Bacillus thuringiensis subsp. thuringiensis inhibits the growth of Yoshida ascites sarcoma both in vivo and in vitro. Exogenous respiration of the tumor cells was unaffected by the protein at a concentration as high as 500 µg/ml. The antitumor protein inhibits the uptake and incorporation of labeled precursors into macromolecules. However, the ratio of incorporation over uptake is not affected by the protein. Further, the protein brings about the leakage of 260-nm-absorbing material, proteins, and 32P-labeled cellular constituents from the Yoshida ascites sarcoma cells. The results show that the action of the antitumor protein appears to alter the cellular permeability of the tumor cells.

Physicochemical, Electrochemical, and Spectroscopic Characterization of Zinc-Based Room-Temperature Molten Electrolytes and Their Application in Rechargeable Batteries

Room-temperature zinc ion-conducting molten electrolytes based on acetamide, urea, and zinc perchlorate or zinc triflate have been prepared and characterized by various physicochemical, spectroscopic, and electrochemical techniques. The ternary molten electrolytes are easy to prepare and can be handled under ambient conditions. They show excellent stability, high ionic conductivity, relatively low viscosity, and other favorable physicochemical and electrochemical properties that make them good electrolytes for rechargeable zinc batteries. Specific conductivities of 3.4 and 0.5 mS cm(-1) at 25 degrees C are obtained for zinc-perchlorate-and zinc-triflate-containing melts, respectively. Vibrational spectroscopic data reveal that the free ion concentration is high in the optimized composition. Rechargeable Zn batteries have been assembled using the molten electrolytes, with gamma-MnO2 as the positive electrode and Zn as the negative electrode. They show excellent electrochemical characteristics with high discharge capacities. This study opens up the possibility of using acetamide-based molten electrolytes as alternate electrolytes in rechargeable zinc batteries. (C) 2009 The Electrochemical Society.

Classification of Nonenzymatic Homologues of Protein Kinases

Protein Kinase-Like Non-kinases (PKLNKs), which are closely related to protein kinases, lack the crucial catalytic aspartate in the catalytic loop, and hence cannot function as protein kinase, have been analysed. Using various sensitive sequence analysis methods, we have recognized 82 PKLNKs from four higher eukaryotic organisms, namely, Homo sapiens, Mus musculus, Rattus norvegicus, and Drosophila melanogaster. On the basis of their domain combination and function, PKLNKs have been classified mainly into four categories: (1) Ligand binding PKLNKs, (2) PKLNKs with extracellular protein-protein interaction domain, (3) PKLNKs involved in dimerization, and (4) PKLNKs with cytoplasmic protein-protein interaction module. While members of the first two classes of PKLNKs have transmembrane domain tethered to the PKLNK domain, members of the other two classes of PKLNKs are cytoplasmic in nature. The current classification scheme hopes to provide a convenient framework to classify the PKLNKs from other eukaryotes which would be helpful in deciphering their roles in cellular processes.