Deletion of cysteine 369 in lysyl hydroxylase 1 eliminates enzyme activity and causes Ehlers-Danlos syndrome type VI.

This study describes the relative contribution of the 10 cysteine residues in lysyl hydroxylase 1 (LH1) to enzyme activity. We have identified a novel mutation of a 15-bp deletion in exon 11 in one LH1 allele, that codes for amino acids 367-371 (DLCRQ), in two unrelated compound heterozygous patients with Ehlers-Danlos type VI. The mutations in their other alleles were a C1119T change (exon 10) and a predicted Q49X (exon 2). We confirmed that the loss of cysteine 369 in the deleted sequence contributed to the diminished enzyme activity by structure/function analysis of mutant LH1 constructs, in which C369 and the nine other cysteines were individually mutated to serine by site-directed mutagenesis of a normal pAcGP67/LH1cDNA construct. Following their expression in an Sf9 insect cell/baculovirus system, SDS-PAGE and Western analysis showed that equivalent levels of correctly-sized (85-kDa) products were secreted. The mutation of residues C369 and also C375, C552 and C687 virtually eliminated LH activity, whereas mutations of C267, C270, and C680 had an intermediate effect. In contrast, the C204S, C484S and C566S constructs had normal activity. Although disulfide bond formation may affect the relative contribution of each cysteine to LH activity, catalytic activity does not appear to be directly related to dimerization of the enzyme.     

A Two-Step Procedure for Obtaining Highly Purified Particulate Guanylate Cyclase From Rat Lung

Abstract

Cuanylate cyclase (CTP pyrophosphate lyase (cyclizing), EC 4.6.1.21, the enzyme catalyzing the formation of cyclic CMP, from GTP, exists in both soluble and membrane-bound forms.1-6 These enzymes have been implicated as key regulatory components in a variety of biological events such as secretion7-10, and smooth muscle relaxation.11-14 Understanding the role these enzymes play in cellular regulation is predicated upon obtaining purified preparations of guanylate cyclase. Several procedures for purifying the soluble enzyme to apparent homogeneity from a variety of tissues have been reported.15-19 The particulate enzyme has been purified to apparent homogeneity from sea urchin sperm20,21 but, to date, homogeneous preparations of particulate guanylate cyclase from mammalian tissues have not been obtained. Here we report our efforts in purifying particulate guanylate cyclase about 8,200- to 52,000-fold from homogenates of rat lung. Portions of this work have appeared in abstract form.     

Antimicrobial activities of Gloriosa superba Linn (Colchicaceae) extracts

The methanol extract of the rhizomes of Gloriosa superba Linn (Colchicaceae) and its subsequent fractions in different solvent systems were screened for antibacterial and antifungal activities. Excellent antifungal sensitivity was expressed by the n-butanol fraction against Candida albicans and Candida glaberata (up to 90%) and against Trichophyton longifusus (78%) followed by the chloroform fraction against Microsporum canis (80%). In the antibacterial bioassay, the crude extract and subsequent fractions showed mild to moderate antibacterial activities. Chloroform fraction displayed highest antibacterial sensitivity against Staphylococcus aureous (88%) followed by the crude extract (59%). The total phenol content of the crude extract and fractions of the plant expressed no significant correlation with the antimicrobial activities.     

Novel Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV2) and Other Coronaviruses: A Genome-wide Comparative Annotation and Analysis

Molecular and Cellular Biochemistry - Novel strain of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV2) causes mild to severe respiratory illness. The early symptoms may be fever, dry...     

Quantitative interpretation of cell rolling velocity distribution

Leukocyte rolling adhesion, facilitated by selectin-mediated interactions, is a highly dynamic process in which cells roll along the endothelial surface of blood vessel walls to reach the site of infection. The most common approach to investigate cell-substrate adhesion is to analyze the cell rolling velocity in response to shear stress changes. It is assumed that changes in rolling velocity indicate changes in adhesion strength. In general, cell rolling velocity is studied at the population level as an average velocity corresponding to given shear stress. However, no statistical investigation has been performed on the instantaneous velocity distribution. In this study, we first developed a method to remove systematic noise and revealed the true velocity distribution to exhibit a log-normal profile. We then demonstrated that the log-normal distribution describes the instantaneous velocity at both the population and single-cell levels across the physiological flow rates. The log-normal parameters capture the cell motion more accurately than the mean and median velocities, which are prone to systematic error. Lastly, we connected the velocity distribution to the molecular adhesion force distribution and showed that the slip-bond regime of the catch-slip behavior of the P-selectin/PSGL-1 interaction is responsible for the variation of cell velocity.     

Effects of sugar feeding supplemented with three plant extracts on some parameters of honey bee colonies

Sugar feeding is crucial to bee colonies during periods without natural nectar resources. The health and the development of bee colonies are affected by the sugar feeding type. Also, some materials can be added to the sugar feeding to boost the ability of bee colonies to withstand parasites. Three materials (mint, cinnamon, and chamomile) are used commonly to control bee parasites (e.g. Varroa mites). In the present study, the effects of these materials on the development and health of bee colonies were assessed. Sugar candy supplemented with these materials plus sugar candy only as a control group were tested. Bee colonies were fed with these feeding types weekly. Then, some parameters were evaluated. The results showed the suitability of the tested feeding types to bee colonies. Building of wax foundations was accelerated in cinnamon group. This group had also the lowest infestation rates with Varroa mites, suggesting a specific role of cinnamon in Varroa control. The colony development was significantly better in chamomile group than the other groups. Mint group showed no variations than the control group in most parameters. All feeding types showed safety to bees based on morphological characteristics and bee survival results. Practically, cinnamon is advised when building of wax combs is required while chamomile is recommended when increasing strength of colonies is needed. The role of cinnamon in controlling Varroa is recommended for further investigations.     

Exploring the non-coding regions in the mtDNA of some honey bee species and subspecies

The sequence of the DNA contains coding and non-coding regions. The role of the non-coding regions is not known and is hypothesized to maintain the structure of the DNA. This study aimed to investigate the structure of the non-coding sequences in honey bees utilizing bioinformatics. The non-coding sequences of the mtDNA of three honey bee species Apis dorosata, Apis florea, Apis cerana, and ten subspecies of Apis mellifera were investigated. Different techniques were utilized to explore the non-coding regions of these bees including sequence analysis, phylogenetic relationships, enzymatic digestion, and statistical tests. Variations in size and sequences of nucleotides were detected in the studied species and subspecies, but with the same nucleotide abundance (i.e. nucleotides A were more than T and nucleotides G were less than C). The phylogenetic tree based on the non-coding regions was partially similar to the known phylogenetic relationships between these bees. The enzymatic digestion using four restriction enzymes confirmed the results of the phylogenetic relationships. The statistical analysis based on numerical codes for nucleotides showed the absence of significant variations between the studied bees in their sequences in a similar way to results of neutrality tests. This study suggests that the non-coding regions have the same functional role in all the studied bees regardless of the number of nucleotides, and not just to maintain the structure of the DNA. This is approximately the first study to shade lights on the non-coding regions of the mtDNA of honey bees.