Analysis of the role of bacterial endospore cortex structure in resistance properties and demonstration of its conservation amongst species

Atrih, Abdelmadjid; Foster, Simon J.

doi:10.1046/j.1365-2672.2001.01394.x

Cited by 73 publications

(73 citation statements)

References 33 publications

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“…Alanine racemase has been reported to be associated with the surface of other Bacillus spores (27). Its presence may be either fortuitous and related to the production of D-alanine needed for cortex synthesis (2,29) or by design and needed for the interconversion of the germinant L-alanine and the germination inhibitor D-alanine (27). In the case of superoxide dismutase, it has been proposed that this enzyme associates with the outer surface of B. subtilis spores and participates in the oxidative cross-linking of outer coat proteins (16,17).…”

Section: Discussionmentioning

confidence: 99%

Identification of the Immunodominant Protein and Other Proteins of theBacillus anthracisExosporium

et al. 2003

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Spores of Bacillus anthracis, the causative agent of anthrax, are enclosed by a prominent loose-fitting, balloon-like layer called the exosporium. Although the exosporium serves as the source of surface antigens and a primary permeability barrier of the spore, its molecular structure and function are not well characterized. In this study, we identified five major proteins in purified B. anthracis (Sterne strain) exosporia. One protein was the recently identified collagen-like glycoprotein BclA, which appears to be a structural component of the exosporium hair-like nap. Using a large panel of unique antispore monoclonal antibodies, we demonstrated that BclA is the immunodominant antigen on the B. anthracis spore surface. We also showed that the BclA protein and not a carbohydrate constituent directs the dominant immune response. In addition, the length of the central (GXX) n repeat region of BclA appears to be strain specific. Two other unique proteins, BxpA and BxpB, were identified. BxpA is unusually rich in Gln and Pro residues and contains several different tandem repeats, which also exhibit strain-specific variation. In addition, BxpA was found to be cleaved approximately in half. BxpB appears to be glycosylated or associated with glycosylated material and is encoded by a gene that (along with bclA) may be part of an exosporium genomic island. The other two proteins identified were alanine racemase and superoxide dismutase, both of which were reported to be associated with the surface of other Bacillus spores. Possible functions of the newly identified proteins are discussed.The genus Bacillus includes a diverse collection of grampositive, rod-shaped, aerobic bacteria that form an endospore (or spore) upon deprivation of an essential nutrient (10, 30). In this process, an asymmetric septation of the starved vegetative cell produces a large and a small genome-containing compartment called the mother cell and forespore, respectively. The mother cell then engulfs the forespore, thereby surrounding it with two opposing cell membranes. A thick layer of modified peptidoglycan called the cortex is synthesized between the two membranes, and proteins synthesized in the mother cell form multiple layers of a spore coat that covers the cortex. While the coat forms the outermost detectable layer for spores of some species (e.g., Bacillus subtilis), in others (e.g., Bacillus anthracis), the spore is enclosed by an additional layer called the exosporium, a prominent, loose-fitting, balloon-like layer also synthesized by the mother cell (12,17). After a final stage of maturation, the mother cell lyses to release the mature spore, which is dormant and capable of persisting in the soil for many years until it encounters a germination signal.Most Bacillus species are not pathogenic to humans. The most notable exception is B. anthracis, the causative agent of anthrax (25). In light of the recent use of B. anthracis spores as a terrorist weapon in the United States and the development of these spores as a weapon of mass destruct...

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Section: Discussionmentioning

confidence: 99%

Identification of the Immunodominant Protein and Other Proteins of theBacillus anthracisExosporium

et al. 2003

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“…Spore dormancy and wet heat resistance are largely dependent on spore core dehydration, which is maintained by a thick layer of modified peptidoglycan (PG) known as the cortex (2,5). While vegetative cell wall PG consists of alternating N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) sugars, approximately 50% of NAM residues in the cortex are converted to muramic-␦-lactam, while an additional portion of the NAM side chains is generally cleaved to a single L-alanine (6)(7)(8)(9)(10)(11).…”

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confidence: 99%

Role of YpeB in Cortex Hydrolysis during Germination of Bacillus anthracis Spores

Bernhards

Popham

2014

J Bacteriol

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The infectious agent of the disease anthrax is the spore of Bacillus anthracis. Bacterial spores are extremely resistant to environmental stresses, which greatly hinders spore decontamination efforts. The spore cortex, a thick layer of modified peptidoglycan, contributes to spore dormancy and resistance by maintaining the low water content of the spore core. The cortex is degraded by germination-specific lytic enzymes (GSLEs) during spore germination, rendering the cells vulnerable to common disinfection techniques. This study investigates the relationship between SleB, a GSLE in B. anthracis, and YpeB, a protein necessary for SleB stability and function. The results indicate that ⌬sleB and ⌬ypeB spores exhibit similar germination phenotypes and that the two proteins have a strict codependency for their incorporation into the dormant spore. In the absence of its partner protein, SleB or YpeB is proteolytically degraded soon after expression during sporulation, rather than escaping the developing spore. The three PepSY domains of YpeB were examined for their roles in the interaction with SleB. YpeB truncation mutants illustrate the necessity of a region beyond the first PepSY domain for SleB stability. Furthermore, site-directed mutagenesis of highly conserved residues within the PepSY domains resulted in germination defects corresponding to reduced levels of both SleB and YpeB in the mutant spores. These results identify residues involved in the stability of both proteins and reiterate their codependent relationship. It is hoped that the study of GSLEs and interacting proteins will lead to the use of GSLEs as targets for efficient activation of spore germination and facilitation of spore cleanup. Bacterial spores from the Bacillus and Clostridium genera are metabolically dormant and are known for their extreme resistance to heat, desiccation, UV radiation, chemicals, and other insults (1-3). These resistance properties allow spores to survive in the environment for extended periods and have made eradication from contaminated sites incredibly difficult (4). Spore dormancy and wet heat resistance are largely dependent on spore core dehydration, which is maintained by a thick layer of modified peptidoglycan (PG) known as the cortex (2, 5). While vegetative cell wall PG consists of alternating N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) sugars, approximately 50% of NAM residues in the cortex are converted to muramic-␦-lactam, while an additional portion of the NAM side chains is generally cleaved to a single L-alanine (6-11).The spore form of Bacillus anthracis is the etiological agent for all types of anthrax infections: inhalational, gastrointestinal, and cutaneous anthrax, as well as the newest form described, injectional anthrax (12, 13). When the spore senses the availability of nutrients, such as when it enters a suitable host, germination is triggered, causing a chain of events that ultimately result in a vegetative cell capable of producing deadly toxins (1, 12). After germinant contact with rec...

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“…The cross-linking occurs between the Dpm on one strand and the D-alanine of a tetrapeptide on an adjacent strand. The spore PG is composed of the inner, germ cell wall layer (10 to 20% of the PG), which resembles the PG of vegetative cells (6,19), and the cortex, which has important and unique chemical modifications (2,3,5,23,28,30). In the cortex, approximately 50% of the muramic acid residues (the residues on every alternate disaccharide) have their peptide side chains completely removed, and these NAM residues are converted to muramic-␦-lactam.…”

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confidence: 99%

Cortex Peptidoglycan Lytic Activity in Germinating Bacillus anthracis Spores

2008

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Bacterial endospore dormancy and resistance properties depend on the relative dehydration of the spore core, which is maintained by the spore membrane and its surrounding cortex peptidoglycan wall. During spore germination, the cortex peptidoglycan is rapidly hydrolyzed by lytic enzymes packaged into the dormant spore. The peptidoglycan structures in both dormant and germinating Bacillus anthracis Sterne spores were analyzed. The B. anthracis dormant spore peptidoglycan was similar to that found in other species. During germination, B. anthracis released peptidoglycan fragments into the surrounding medium more quickly than some other species. A major lytic enzymatic activity was a glucosaminidase, probably YaaH, that cleaved between Nacetylglucosamine and muramic-␦-lactam. An epimerase activity previously proposed to function on spore peptidoglycan was not detected, and it is proposed that glucosaminidase products were previously misidentified as epimerase products. Spore cortex lytic enzymes and their regulators are attractive targets for development of germination inhibitors to kill spores and for development of activators to cause loss of resistance properties for decontamination of spore release sites.Bacterial endospores are metabolically dormant and are highly resistant to many treatments that rapidly kill vegetative cells. The resistance properties are dependent on the relative dehydration of the spore core or cytoplasm and the high concentrations of calcium dipicolinic acid (DPA) and other core solutes (26). When exposed to a nutrient-rich environment, spores initiate germination and outgrowth into vegetative cells. Nutrient germinant molecules bind to Ger protein family receptors in the inner forespore membrane surrounding the spore core and trigger biophysical changes (20). The spore becomes partially rehydrated as water enters the spore core, and the calcium DPA that was in the core is released into the environment. To complete the germination process and resume metabolism (11, 24), the spore must go through a second stage, which is more biochemical (20). This involves activation of spore cortex peptidoglycan (PG) lytic enzymes, which are present in an inactive state within the dormant spore. As the cortex is degraded, the cell expands to its full size and regains full metabolic activity. Due to the key role that cortex lytic enzymes play in germination, they are potential targets for spore decontamination methods that could involve either irreversible inactivation or gratuitous activation of the enzymes. The latter would lead to premature spore germination into vegetative forms that are much more easily killed.PG consists of glycan strands with alternating residues of N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) that are cross-linked by peptide side chains located on the NAM residues (Fig. 1). While the PG structure in vegetative cells can vary between species, primarily in the amino acid composition of the peptide side chains, spore PG structure has been found to be highly conserved acro...

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Analysis of the role of bacterial endospore cortex structure in resistance properties and demonstration of its conservation amongst species

Cited by 73 publications

References 33 publications

Identification of the Immunodominant Protein and Other Proteins of theBacillus anthracisExosporium

Identification of the Immunodominant Protein and Other Proteins of theBacillus anthracisExosporium

Role of YpeB in Cortex Hydrolysis during Germination of Bacillus anthracis Spores

Cortex Peptidoglycan Lytic Activity in Germinating Bacillus anthracis Spores

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