The diacetamidodideoxyuronic-acid-containing glycan chain of Bacillus stearothermophilus NRS 2004/3a represents the secondary cell-wall polymer of wild-type B. stearothermophilus strains

ABSTRACT Two Bacillus stearothermophilus wild-type strains were investigated regarding a common recognition and binding mechanism between the S-layer protein and the underlying cell envelope layer. The S-layer protein from B. stearothermophilusPV72/p6 has a molecular weight of 130,000 and assembles into a hexagonally ordered lattice. The S-layer from B. stearothermophilus ATCC 12980 shows oblique lattice symmetry and is composed of subunits with a molecular weight of 122,000. Immunoblotting, peptide mapping, N-terminal sequencing of the whole S-layer protein from B. stearothermophilus ATCC 12980 and of proteolytic cleavage fragments, and comparison with the S-layer protein from B. stearothermophilus PV72/p6 revealed that the two S-layer proteins have identical N-terminal regions but no other extended structurally homologous domains. In contrast to the heterogeneity observed for the S-layer proteins, the secondary cell wall polymer isolated from peptidoglycan-containing sacculi of the different strains showed identical chemical compositions and comparable molecular weights. The S-layer proteins could bind and recrystallize into the appropriate lattice type on native peptidoglycan-containing sacculi from both organisms but not on those extracted with hydrofluoric acid, leading to peptidoglycan of the A1γ chemotype. Affinity studies showed that only proteolytic cleavage fragments possessing the complete N terminus of the mature S-layer proteins recognized native peptidoglycan-containing sacculi as binding sites or could associate with the isolated secondary cell wall polymer, while proteolytic cleavage fragments missing the N-terminal region remained unbound. From the results obtained in this study, it can be concluded that S-layer proteins from B. stearothermophilus wild-type strains possess an identical N-terminal region which is responsible for anchoring the S-layer subunits to a secondary cell wall polymer of identical chemical composition.

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TOWARDS THE SYNTHESIS OF THE SECONDARY CELL WALL POLYMER (SCWP) OF BACILLUS STEAROTHERMOPHILUS STRAIN PV72/p6

XXIst International Carbohydrate Symposium 2002 ◽

10.1100/tsw.2002.555 ◽

2002 ◽

Author(s):

A. G. Hedenetz ◽

B. Schwarz ◽

P. Olf ◽

F. M. Unger ◽

W. Schmid

Keyword(s):

Cell Wall ◽

Secondary Cell Wall ◽

Bacillus Stearothermophilus ◽

Cell Wall Polymer ◽

Secondary Cell Wall Polymer

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Influence of the Secondary Cell Wall Polymer on the Reassembly, Recrystallization, and Stability Properties of the S-Layer Protein from Bacillus stearothermophilus PV72/p2

Journal of Bacteriology ◽

10.1128/jb.180.16.4146-4153.1998 ◽

1998 ◽

Vol 180 (16) ◽

pp. 4146-4153 ◽

Cited By ~ 37

Author(s):

Margit Sára ◽

Christine Dekitsch ◽

Harald F. Mayer ◽

Eva M. Egelseer ◽

Uwe B. Sleytr

Keyword(s):

Cell Wall ◽

Self Assembly ◽

Secondary Cell Wall ◽

Bacillus Stearothermophilus ◽

Guanidine Hydrochloride ◽

Terminal Region ◽

Water Soluble ◽

Outer Face ◽

Cell Wall Polymer ◽

Secondary Cell Wall Polymer

ABSTRACT The high-molecular-weight secondary cell wall polymer (SCWP) fromBacillus stearothermophilus PV72/p2 is mainly composed ofN-acetylglucosamine (GlcNAc) andN-acetylmannosamine (ManNAc) and is involved in anchoring the S-layer protein via its N-terminal region to the rigid cell wall layer. In addition to this binding function, the SCWP was found to inhibit the formation of self-assembly products during dialysis of the guanidine hydrochloride (GHCl)-extracted S-layer protein. The degree of assembly (DA; percent assembled from total S-layer protein) that could be achieved strongly depended on the amount of SCWP added to the GHCl-extracted S-layer protein and decreased from 90 to 10% when the concentration of the SCWP was increased from 10 to 120 μg/mg of S-layer protein. The SCWP kept the S-layer protein in the water-soluble state and favored its recrystallization on solid supports such as poly-l-lysine-coated electron microscopy grids. Derived from the orientation of the base vectors of the oblique S-layer lattice, the subunits had bound with their charge-neutral outer face, leaving the N-terminal region with the polymer binding domain exposed to the ambient environment. From cell wall fragments about half of the S-layer protein could be extracted with 1 M GlcNAc, indicating that the linkage type between the S-layer protein and the SCWP could be related to that of the lectin-polysaccharide type. Interestingly, GlcNAc had an effect on the in vitro self-assembly and recrystallization properties of the S-layer protein that was similar to that of the isolated SCWP. The SCWP generally enhanced the stability of the S-layer protein against endoproteinase Glu-C attack and specifically protected a potential cleavage site in position 138 of the mature S-layer protein.

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Evidence that the N-terminal part of the S-layer protein from Bacillus stearothermophilus PV72/p2 recognizes a secondary cell wall polymer.

Journal of Bacteriology ◽

10.1128/jb.179.12.3892-3898.1997 ◽

1997 ◽

Vol 179 (12) ◽

pp. 3892-3898 ◽

Cited By ~ 95

Author(s):

W Ries ◽

C Hotzy ◽

I Schocher ◽

U B Sleytr ◽

M Sára

Keyword(s):

Cell Wall ◽

Secondary Cell Wall ◽

Bacillus Stearothermophilus ◽

Terminal Part ◽

Cell Wall Polymer ◽

Secondary Cell Wall Polymer

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Identification of Two Binding Domains, One for Peptidoglycan and Another for a Secondary Cell Wall Polymer, on the N-Terminal Part of the S-Layer Protein SbsB from Bacillus stearothermophilus PV72/p2

Journal of Bacteriology ◽

10.1128/jb.180.24.6780-6783.1998 ◽

1998 ◽

Vol 180 (24) ◽

pp. 6780-6783 ◽

Cited By ~ 36

Author(s):

Margit Sára ◽

Eva M. Egelseer ◽

Christine Dekitsch ◽

Uwe B. Sleytr

Keyword(s):

Amino Acids ◽

Cell Wall ◽

Secondary Cell Wall ◽

Bacillus Stearothermophilus ◽

Binding Domain ◽

Terminal Part ◽

Binding Domains ◽

Cell Wall Polymer ◽

Function Relationship ◽

Secondary Cell Wall Polymer

ABSTRACT First studies on the structure-function relationship of the S-layer protein from B. stearothermophilus PV72/p2 revealed the coexistence of two binding domains on its N-terminal part, one for peptidoglycan and another for a secondary cell wall polymer (SCWP). The peptidoglycan binding domain is located between amino acids 1 to 138 of the mature S-layer protein comprising a typical S-layer homologous domain. The SCWP binding domain lies between amino acids 240 to 331 and possesses a high serine plus glycine content.

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High-Affinity Interaction between the S-Layer Protein SbsC and the Secondary Cell Wall Polymer of Geobacillus stearothermophilus ATCC 12980 Determined by Surface Plasmon Resonance Technology

Journal of Bacteriology ◽

10.1128/jb.00294-07 ◽

2007 ◽

Vol 189 (19) ◽

pp. 7154-7158 ◽

Cited By ~ 19

Author(s):

Judith Ferner-Ortner ◽

Christoph Mader ◽

Nicola Ilk ◽

Uwe B. Sleytr ◽

Eva M. Egelseer

Keyword(s):

Amino Acids ◽

Cell Wall ◽

Surface Plasmon Resonance ◽

Surface Plasmon ◽

Plasmon Resonance ◽

Secondary Cell Wall ◽

Geobacillus Stearothermophilus ◽

Binding Behavior ◽

Cell Wall Polymer ◽

Secondary Cell Wall Polymer

ABSTRACT Surface plasmon resonance studies using C-terminal truncation forms of the S-layer protein SbsC (recombinant SbsC consisting of amino acids 31 to 270 [rSbsC31-270] and rSbsC31-443) and the secondary cell wall polymer (SCWP) isolated from Geobacillus stearothermophilus ATCC 12980 confirmed the exclusive responsibility of the N-terminal region comprising amino acids 31 to 270 for SCWP binding. Quantitative analyses indicated binding behavior demonstrating low, medium, and high affinities.

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Structural and Functional Analyses of the Secondary Cell Wall Polymer of Bacillus sphaericus CCM 2177 That Serves as an S-Layer-Specific Anchor

Journal of Bacteriology ◽

10.1128/jb.181.24.7643-7646.1999 ◽

1999 ◽

Vol 181 (24) ◽

pp. 7643-7646 ◽

Cited By ~ 78

Author(s):

Nicola Ilk ◽

Paul Kosma ◽

Michael Puchberger ◽

Eva M. Egelseer ◽

Harald F. Mayer ◽

...

Keyword(s):

Cell Wall ◽

Hydrofluoric Acid ◽

Secondary Cell Wall ◽

Bacillus Sphaericus ◽

Nuclear Magnetic Resonance Analysis ◽

Terminal Part ◽

Resonance Analysis ◽

Cell Wall Polymer ◽

Functional Analyses ◽

Secondary Cell Wall Polymer

ABSTRACT Sacculi of Bacillus sphaericus CCM 2177 contain a secondary cell wall polymer which was completely extracted with 48% hydrofluoric acid. Nuclear magnetic resonance analysis showed that the polymer is composed of repeating units, as follows: →3)-[4,6-O-(1-carboxyethylidene)]∼0.5-β-d-ManpNAc-(1→4)-β-d-GlcpNAc-(1→. The N-terminal part of the S-layer protein carrying S-layer homologous motifs recognizes this polymer as a binding site.

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