The sequence of a promoter determines not only the efficiency with which it forms complex RNA polymerase, but also concentration nucleoside triphosphate (NTP) required for initiating transcription. Escherichia coli ribosomal ( rrn P1) promoters require high NTP concentrations efficient transcription because they form unusually short-lived complexes polymerase; [adenosine or guanosine (ATP GTP), depending on P1 promoter] are needed to bind and stabilize open complex. ATP GTP concentrations,...

Spores of Bacillus anthracis, the causative agent anthrax, are enclosed by a prominent loose-fitting, balloon-like layer called exosporium. Although exosporium serves as source surface antigens and primary permeability barrier spore, its molecular structure function not well characterized. In this study, we identified five major proteins in purified B. anthracis (Sterne strain) exosporia. One protein was recently collagen-like glycoprotein BclA, which appears to be structural component...

Journal Article Rapid confirmation of single copy lambda prophage integration by PCR Get access Bradford S. Powell, Powell 1Laboratory Chromosome Biology, ABL-Basic Research Program, NCI-Frederick Cancer and Development CenterFrederick, MD 21702-12013Department Tumor The Institute Medical Science, University TokyoPO Takanawa 108, Japan Search for other works this author on: Oxford Academic PubMed Google Scholar Donald L. Court, Court * 21702-1201 *To whom correspondence should be addressed...

Spores of Bacillus anthracis, the causative agent anthrax, are enclosed by a prominent loose fitting layer called exosporium. The exosporium consists basal and an external hairlike nap. filaments nap composed highly immunogenic glycoprotein BclA, which has long, central collagen-like region with multiple XXG repeats. Most triplet repeats PTG, nearly all contain threonine residue, providing potential sites for O-glycosylation. In this study, we demonstrated that two O-linked oligosaccharides,...

Although quinolones are the most commonly prescribed antibacterials, their use is threatened by an increasing prevalence of resistance. The common causes quinolone resistance mutations a specific serine or acidic residue in A subunit gyrase topoisomerase IV. These amino acids proposed to serve as critical enzyme-quinolone interaction site anchoring water-metal ion bridge that coordinates drug binding. To probe role bridge, we characterized wild-type, GrlA E85K , S81F/E85K E85A S81F/E85A and...

Adaptive immunity in jawless vertebrates (lamprey and hagfish) is mediated by lymphocytes that undergo combinatorial assembly of leucine-rich repeat (LRR) gene segments to create a diverse repertoire variable lymphocyte receptor (VLR) genes. Immunization with particulate antigens induces VLR-B-bearing secrete antigen-specific VLR-B antibodies. Here, we describe the production recombinant antibodies specific for BclA, major coat protein Bacillus anthracis spores. The possess 8-10 uniform...

Although bacterial gyrase and topoisomerase IV have critical interactions with positively supercoiled DNA, little is known about the actions of these enzymes on overwound substrates. Therefore, abilities Bacillus anthracis Escherichia coli to relax cleave DNA were analyzed. Gyrase removed positive supercoils ∼10-fold more rapidly processively than it introduced negative into relaxed DNA. In time-resolved single-molecule measurements, burst rates ∼100 per second (average size was 6.2...

The pyrBI operon of Escherichia coli K-12 encodes the subunits pyrimidine biosynthetic enzyme aspartate transcarbamylase (carbamoylphosphate:L-aspartate carbamoyltransferase, EC 2.1.3.2). Expression this apparently is negatively regulated by intracellular levels UTP. To elucidate regulatory mechanism in which UTP functions, nucleotide sequence promoter-regulatory region was determined and DNA fragments containing were transcribed vitro. These experiments revealed a rho-independent...

Bacillus anthracis spores, which cause anthrax, are enclosed by an exosporium consisting of a basal layer and external hair-like nap. The filaments the nap composed BclA, glycoprotein containing distinct N-terminal (NTD) C-terminal (CTD) domains separated extended collagen-like central region. In this study, we used immunogold electron microscopy to show that CTD BclA forms distal end each filament nap, indicating NTD is attached layer. Ten randomly chosen anti-BclA monoclonal antibodies,...

Bacillus anthracis, the causative agent of anthrax, is considered a serious threat as bioweapon. The drugs most commonly used to treat anthrax are quinolones, which act by increasing levels DNA cleavage mediated topoisomerase IV and gyrase. Quinolone resistance often associated with specific serine mutations in these enzymes. Therefore, determine basis for quinolone action resistance, we characterized wild-type B. anthracis IV, GrlA(S81F) GrlA(S81Y) quinolone-resistant mutants, effects...

Bacillus anthracis spores, the cause of anthrax, are enclosed by a prominent loose-fitting structure called exosporium. The exosporium is composed basal layer and an external hair-like nap. filaments nap apparently formed single collagen-like glycoprotein BclA, whereas several different proteins form or tightly associated with layer. In this study, we used immunogold electron microscopy to demonstrate that BxpB (also ExsF) component Binding anti-BxpB monoclonal antibody was greatly increased...

Anthrax, a disease caused by Bacillus anthracis, affects animals and humans. Because the inert spore is infectious form of organism that first contacts potential host, interaction between host exosporium vital to initiation disease. Here, we demonstrate integrin Mac-1 essential for recognition major protein BclA phagocytic cells. Expression Mac-1, but not p150/95, in CHO cells markedly enhanced infection with Sterne strain B. anthracis spores (WT spores). Conversely, CD11b(-/-) macrophages...

One of the nonspecific defense mechanisms higher animals is their ability to limit iron availability infecting bacteria. Thus it has been argued that all pathogenic bacteria must have special obtain in host environment. Salmonella typhimurium known produce a siderophore, enterobactin, with which can from transferrin. Previous studies indicated production this molecule necessary for intraperitoneally injected. S. cells cause mouse typhoid, largely intracellular infection. We reexamined...

Anthrax, a potentially lethal disease of animals and humans, is caused by the Gram-positive spore-forming bacterium Bacillus anthracis. The outermost exosporium layer B. anthracis spores contains an external hair-like nap formed glycoprotein BclA. Recognition BclA integrin Mac-1 promotes spore uptake professional phagocytes, resulting in carriage to sites germination bacterial growth distant lymphoid organs. We show that CD14 binds rhamnose residues acts as coreceptor for binding Mac-1. In...

Summary Spores of Bacillus anthracis are enclosed by an exosporium composed a basal layer and external hair‐like nap. The nap is formed collagen‐like glycoprotein called BclA, while the contains many different proteins, one which spore‐specific alanine racemase (Alr). In this study, we employed fluorescence microscopy fluorescently labelled anti‐Alr monoclonal antibody (mAb) to examine distribution Alr within exosporium. Binding mAb occurred over approximately three‐quarters but not in...

Quinolones, which target gyrase and topoisomerase IV, are the most widely prescribed antibacterials worldwide. Unfortunately, their use is threatened by increasing prevalence of target-mediated drug resistance. Greater than 90% mutations that confer quinolone resistance act disrupting enzyme–drug interactions coordinated a critical water–metal ion bridge. Quinazolinediones quinolone-like drugs but lack skeletal features necessary to support bridge interaction. These compounds clinical...

Recent use of Bacillus anthracis spores as a bioweapon has highlighted the need for continuous monitoring system. Current systems rely on antibody-derived probes, which are not hardy enough to withstand long-term under extreme conditions. We describe new, phage-derived probes that can be used robust substitutes antibodies.From landscape phage library with random octapeptides displayed all copies major coat protein fd-tet, we selected clones bound B. (Sterne strain). ELISA, micropanning, and...

ABSTRACT Currently available detectors for spores of Bacillus anthracis , the causative agent anthrax, are inadequate frontline use and general monitoring. There is a critical need simple, rugged, inexpensive capable accurate direct identification B. spores. Necessary components in such stable ligands that bind tightly specifically to target By screening phage display peptide library, we identified family peptides, with consensus sequence TYPXPXR, selectively We extended this work by...

Expression of the pyrBI operon Escherichia coli K-12, which encodes subunits pyrimidine biosynthetic enzyme aspartate transcarbamylase, is negatively regulated by intracellular levels UTP. Previous experiments suggested a unique model for regulation expression in low UTP cause close coupling transcription and translation leader region. This suppresses transcriptional termination at an attenuator preceding structural genes. In this study, we examined regulatory role attenuation expression. To...

Pyrimidine-mediated regulation of pyrBI operon expression in Escherichia coli K-12 occurs primarily by an attenuation control mechanism. Previous studies have suggested a model for which low intracellular levels UTP cause close coupling transcription and translation within the leader region. This apparently prevents transcriptional termination at attenuator (a rho-independent terminator) located 23 base pairs before structural genes open reading frame 44-amino acid polypeptide. Presumably,...

The outermost layer of the Bacillus anthracis spore is exosporium, which composed a paracrystalline basal and an external hair-like nap. filaments nap are formed by collagen-like glycoprotein called BclA, while contains several different proteins. One putative proteins ExsY. In this study, we constructed DeltaexsY mutant B. anthracis, devoid ExsY, examined assembly exosporium on spores produced strain. Our results show that aberrant, with arrested after formation cap-like fragment covers one...

Spores of Bacillus anthracis are enclosed by an exosporium composed a basal layer and external hair-like nap. The nap is apparently formed single glycoprotein, while the contains many different structural proteins several enzymes. One enzymes Alr, alanine racemase capable converting spore germinant l-alanine to germination inhibitor d-alanine. Unlike other characterized proteins, Alr nonuniformly distributed in might have second location. In this study, we demonstrated that expression alr...