Partial assignment of disulfide pairs in neurophysins

The original report assigning the pairing of neurophysin's 14 half-cystine residues (Schlesinger et al. (1972), Proc. Natl. Acad. Sci., U.S.A., 69,3350-3353) was based on an incorrect amino acid sequence. In the present study, re-investigation of the results of proteolytic fragmentation of bovine neurophysins indicates that the majority of the original assignments were incorrect. Three disulfide pairs are now assigned as Cys21-Cys44, Cys67-Cys85 and Cys74-Cys79. The pairing pattern indicates that neurophysin's variable carboxyl terminal region, separately encoded by the third gene exon, does not form a self-contained domain.     

Effects of replacement of active site residue glutamine 231 on activity and allosteric properties of aspartate transcarbamoylase.

Since crystallographic studies on Escherichia coli aspartate transcarbamoylase (ATCase) indicate that Gln 231 is in the active site of the enzyme and participates in the binding of the substrate, aspartate, it seemed of interest to examine mutant enzymes in which Gln 231 was replaced by Asn or Ile. The two mutant forms containing amino acid substitutions were characterized by a combination of steady-state kinetics, hydrodynamic measurements, and equilibrium ligand binding techniques. Both mutant forms exhibited a dramatic reduction in the affinity of the protein for substrates and substrate analogues as well as a very large decrease in catalytic activity. Moreover, the amino acid substitutions introduced within the active site of the enzyme led to unusual allosteric properties in the mutant enzymes. Although the bisubstrate analogue N-(phosphonoacetyl)-L-aspartate promotes the characteristic global conformational change in the mutant forms that is observed with the wild-type enzyme, the combination of substrate and substrate analogue does not. Cooperativity with respect to substrate binding is largely reduced compared to wild-type ATCase. Also, the effector molecules ATP and CTP which bind to the regulatory chains have dramatic effects on the activity of the mutant enzymes containing replacements for Gln 231 in the catalytic chains. In stark contrast to the wild-type enzyme, in which effects of nucleotides are manifested primarily by changes in the K0.5 of the enzyme, ATP and CTP have large effects on the Vmax of the mutant enzymes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Removal of endotoxin from antibody preparations for clinical use

Cell Biochemistry and Biophysics - Despite attempts to maintain asepsis, good manufacturing practices, and the use of terminal sterilization by millipore filtration, the nuclear practitioner is...     

Passage number of cancer cell lines: Importance,intricacies, and way-forward

Cancer cell lines play a crucial role as invaluable models in cancer research, facilitating the examination of cancer progression as well as the advancement of diagnostics and treatments. While they may not perfectly replicate the original tumor, they generally exhibit similar characteristics. Low-passage cancer cell lines are generally preferred due to their closer resemblance to the original tumor, as long-term culturing can alter the genetic and molecular profiles of a cell line thereby highlighting the importance of monitoring the passage number (PN). Variations in proliferation, migration, gene expression, and drug sensitivity can be linked to PN differences. PN can also influence DNA methylation levels, metabolic profiles, and the expression of genes/or proteins in cancer cell lines. When conducting research on cancer cell lines, it is crucial for researchers to carefully select the appropriate PN to maintain consistency and reliability of results. Moreover, to ensure dependability and replicability, scientists ought to actively track the growth, migration, and gene/or protein profiles of cancer cell lines at specific PNs. This approach enables the identification of the most suitable range of PNs for experiments, guaranteeing consistent and precise results. Additionally, such efforts serve to minimize disparities and uphold the integrity of research. In this review, we have laid out recommendations for laboratories to overcome these PN discrepancies when working with cancer cell lines.     

Expression of R-plasmid functions during anaerobic growth of an Escherichia coli K-12 host.

The normal habitat of enteric bacteria is largely anaerobic. Expression of the three characteristic properties of R-plasmids, drug resistance, vegetative replication, and fertility, was therefore studied in Escherichia coli K-12 during anaerobiosis. Replication and drug resistance functions were not altered in the 45 R-plasmids studies, whereas the expression of fertility varied considerably among different R-plasmids during anaerobiosis. The R-plasmids could be divided into three groups, one showing a strong, the second a moderate, and the third little or no reduction of fertility by anaerobiosis. Plasmid-determined sensitivity to F-, N-, and I-specific phage, respectively, was well, although not absolutely, correlated with each of three groups mentioned. Anaerobiosis-aerobiosis appears to change the fertility of type F R-plasmids by influencing the degree of repression of their fertility functions such as the formation of sex pili. Although the minimum inhibitory concentrations of ampicillin, chloramphenicol, streptomycin, and tetracycline were unaltered by anaerobiosis, sulfonamide was found to be four- to eightfold less active under this condition in both resistant and sensitive strains. A surprisingly high frequency and uniformity of minimal inhibitory concentrations was observed for R-plasmid-mediated resistance to streptomycin and chloramphenicol.     

Three new species of thrasya (Gramineae) from Brazil

Thrasya oreophila, T. glaziovii, andT. auricoma are described, discussed, and illustrated.     

Composite response of naive T cells to stimulation with the autologous lymphoblastoid cell line is mediated by CD4 cytotoxic T cell clones and includes an Epstein-Barr virus-specific component.

We have approached the challenge of generating a primary T cell response to Epstein-Barr virus (EBV) in vitro by stimulating naive T cells with the autologous EBV-transformed lymphoblastoid cell line (LCL), a rich source of EBV-associated cytotoxic T lymphocyte (CTL) epitopes. Responsive T cells from three EBV-seronegative donors were cloned in agarose, phenotyped for T cell markers by flow cytometry, and their cytotoxic properties analyzed in the 51Cr release assay. Most clones (greater than 95%) expressed the CD4 phenotype and 59% of these clones showed cytotoxic properties. The dominant CTL response was specific for FCS-associated epitopes presented by FCS-grown autologous LCL target cells and was restricted by class II HLA antigens. Other clonal components included: (i) an EBV-specific response by HLA-restricted CD4 CTL clones that did not discriminate between A- and B-type EBV transformants; (ii) an EBV-specific response by an HLA-restricted CD4 CTL clone that discriminated between A- and B-type transformants, and (iii) a nonspecific cytotoxic response by CD3+,4+,8-, CD3+,4-,8-, and CD3-,4-,8- clones that were broadly allotypic or restricted to the lysis of K562 target cells. The EBV-specific CTL clones did not lyse the autologous EBV-negative B or T cell blasts and their specificity patterns of lysis were supported by the cold target competition data. These studies highlight the role of CD4 CTL in the establishment in vitro of a primary immune response to a human virus.     

Treatment of periodontal disease using xanthan based chlorhexidine gel

People of all ages are suffering from periodontal disease. It causes indirect damage in the oral cavity. It is of interest to evaluate the efficacy of xanthan-based chlorhexidine gel (Xan-CHX) in patients with mild-severe chronic periodontitis. Five patients with 60 sites were divided in two groups. Group A (treated with SRP) and group B (treated with Chlosite i.e., SRP + CHL). The recorded clinical parameters were Plaque index (PI), Gingival index (GI), Bleeding index (BI), and Clinical attachment Level (CAL) with sub gingival plaque subjected to microbial analysis. Significant reduction was observed in both groups. However, group B (treated with Chlosite i.e., SRP + CHL) showed statistically significant improvement on above mentioned parameters as compared to group A. Data suggest that in the treatment of periodontal disease (viz. PI, GI, BI and CAL) combination of SRP and Chlosite showed added benefits over only SRP.