The Bacillus subtilis 168 csn gene encodes a chitosanase with similar properties to a Streptomyces enzyme

Rivas, Luis A.; Parro, Vı́ctor; Moreno‐Paz, Mercedes; Mellado, Rafael P.

doi:10.1099/00221287-146-11-2929

Cited by 34 publications

(22 citation statements)

References 40 publications

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“…Remarkably, most of these proteins are normally considered to be intracellular cytoplasmic proteins. Similar to Est55, no N-terminal cleavage can be detected for any of these proteins, as judged from sequence comparison, except for chitosanase (band i) which had lost its signal peptide; secretion of chitosanase is SecA dependent (33).…”

Section: Resultsmentioning

confidence: 95%

Nonclassical Protein Secretion by Bacillus subtilis in the Stationary Phase Is Not Due to Cell Lysis

Yang¹,

Ewis²,

Zhang³

et al. 2011

Journal of Bacteriology

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The carboxylesterase Est55 has been cloned and expressed in Bacillus subtilis strains. Est55, which lacks a classical, cleavable N-terminal signal sequence, was found to be secreted during the stationary phase of growth such that there is more Est55 in the medium than inside the cells. Several cytoplasmic proteins were also secreted in large amounts during late stationary phase, indicating that secretion in B. subtilis is not unique to Est55. These proteins, which all have defined cytoplasmic functions, include GroEL, DnaK, enolase, pyruvate dehydrogenase subunits PdhB and PdhD, and SodA. The release of Est55 and those proteins into the growth medium is not due to gross cell lysis, a conclusion that is supported by several lines of evidence: constant cell density and secretion in the presence of chloramphenicol, constant viability count, the absence of EF-Tu and SecA in the culture medium, and the lack of effect of autolysin-deficient mutants. The shedding of these proteins by membrane vesicles into the medium is minimal. More importantly, we have identified a hydrophobic ␣-helical domain within enolase that contributes to its secretion. Thus, upon the genetic deletion or replacement of a potential membrane-embedding domain, the secretion of plasmid gene-encoded mutant enolase is totally blocked, while the wild-type chromosomal enolase is secreted normally in the same cultures during the stationary phase, indicating differential specificity. We conclude that the secretion of Est55 and several cytoplasmic proteins without signal peptides in B. subtilis is a general phenomenon and is not a consequence of cell lysis or membrane shedding; instead, their secretion is through a process(es) in which protein domain structure plays a contributing factor.Bacillus subtilis secretes large amounts of proteins into the growth medium (43). Of the known secretory pathways in B. subtilis, the majority of proteins are exported from the cytoplasm by the Sec-dependent pathway, through which secretory proteins are synthesized as precursors with typical cleavable N-terminal signal peptides (3, 36). Fewer proteins are released into the medium via the cleavable twin-arginine translocation (TAT) system (39). Still other proteins are exported into the medium via ATP-binding cassette transporters, a dedicated pseudopilin export pathway, a competence development system or an ESAT-6 (Mycobacterium tuberculosis early secreted antigenic target of 6 kDa)-like system (31).The genome of B. subtilis 168 is 4,215 kbp in length and contains about 4,100 genes that are predicted to include over 250 extracellular proteins; the majority of these proteins are secreted through the aforementioned pathways (3, 18). However, proteomic studies have revealed that genome-based predictions reflect only 50% of the actual composition of the extracellular proteome. This significant discrepancy is mainly due to the difficulties in the prediction of extracellular proteins lacking signal peptides (including cytoplasmic proteins) and lipoproteins (3, 18). These findin...

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Section: Resultsmentioning

confidence: 95%

Nonclassical Protein Secretion by Bacillus subtilis in the Stationary Phase Is Not Due to Cell Lysis

Yang¹,

Ewis²,

Zhang³

et al. 2011

Journal of Bacteriology

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“…A detailed study of chitosanase expression in the absence of chitosan was performed on B. subtilis 168. 11 The production of the GH46 chitosanase by this strain was not induced by the presence of chitosan but was rather temporally regulated, as the transcription of its gene was maximal at the transition period between exponential and stationary phase of the culture and was not subjected to catabolite repression in the presence of glucose. This kind of transcriptional behavior raises questions about the function of chitosanase in Bacillus physiology, suggesting possible roles in stress response or differentiation rather than the intuitively simple role in chitosan metabolism.…”

Section: Ryszard Brzezinskimentioning

confidence: 99%

Uncoupling chitosanase production from chitosan

Brzeziński¹

2011

Bioengineered Bugs

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“…Plasmids pWQ, pXQ, pYQ, and pZQ, used in the in vivo assay, were constructed by inserting a pSN40 (Parro and Mellado 1998) derived fragment containing the sipW, sipX, sipY or sipZ gene into pQC101, a pQC10 (Rivas et al 2000) derivative encoding pre-chitosanase (pre-Csn). All sip genes were placed under the control of the lac promoter, whilst csn was transcribed from its own promoter.…”

Section: Nick Geukens · Victor Parro · Luis a Rivas · Rafael P Mellmentioning

confidence: 99%

Functional analysis of the Streptomyces lividans type I signal peptidases

Parro

Rivas

Mellado

et al. 2001

Archives of Microbiology

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Type I signal peptidases are responsible for the proteolytic cleavage of the signal peptide of secreted proteins. In the gram-positive bacterium Streptomyces lividans, four adjacent genes (sipW, sipX, sipY and sipZ) were isolated encoding putative type I signal peptidases. In this work, the different sip genes were cloned and expressed. Subsequently, the Sip proteins were purified to raise antibodies. Although the four Sip proteins share a low degree of sequence similarity and differ significantly in size and pI, anti-Sip antibodies cross-reacted intensively. Functional signal peptidase processing activity for each of these Sip proteins was shown both in vitro and in vivo. The different Sip proteins did not exhibit the same cleavage efficiency on the Bacillus subtilis pre-chitosanase.

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The Bacillus subtilis 168 csn gene encodes a chitosanase with similar properties to a Streptomyces enzyme

Cited by 34 publications

References 40 publications

Nonclassical Protein Secretion by Bacillus subtilis in the Stationary Phase Is Not Due to Cell Lysis

Nonclassical Protein Secretion by Bacillus subtilis in the Stationary Phase Is Not Due to Cell Lysis

Uncoupling chitosanase production from chitosan

Functional analysis of the Streptomyces lividans type I signal peptidases

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