Surface Of Bacterial Cell Wall Research Articles

A mechanistic analysis of bacterial recognition and serine protease cascade initiation in larval hemolymph of Manduca sexta

Serine protease cascades have evolved in vertebrates and invertebrates to mediate rapid defense responses. Previous biochemical studies showed that in hemolymph of a caterpillar, Manduca sexta, recognition of fungi by β-1,3-glucan recognition proteins (βGRP1 and βGRP2) or recognition of bacteria by peptidoglycan recognition protein-1 (PGRP1) and microbe binding protein (MBP) results in autoactivation of hemolymph protease-14 precursor (proHP14). HP14 then activates downstream members of a protease cascade leading to the melanization immune response. ProHP14 has a complex domain architecture, with five low-density lipoprotein receptor class A repeats at its amino terminus, followed by a Sushi domain, a Sushi domain variant called Wonton, and a carboxyl-terminal serine protease catalytic domain. Its zymogen form is activated by specific proteolytic cleavage at the amino-terminal end of the protease domain. While a molecular mechanism for recognition and triggering the response to β-1,3-glucan has been delineated, it is unclear how bacterial recognition stimulates proHP14 activation. To fill this knowledge gap, we expressed the two domains of M. sexta MBP and found that the amino-terminal domain binds to diaminopimelic acid-peptidoglycan (DAP-PG). ProHP14 bound to both the carboxyl-terminal domain (MBP-C) and amino-terminal domain (MBP-N) of MBP. In the mixture of DAP-PG, MBP, and larval plasma, inclusion of an HP14 fragment composed of LDLa repeats 2–5 (LDLa2–5) or MBP-C significantly reduced prophenoloxidase activation, likely by competing with the interactions of the full-length proteins, and suggesting that molecular interactions involving these regions of proHP14 and MBP take part in proHP14 activation in response to peptidoglycan. Using a series of N-terminally truncated versions of proHP14, we found that autoactivation required LDLa2–5. The optimal ratio of PGRP1, MBP, and proHP14 is close to 3:2:1. In summary, proHP14 autoactivation by DAP-type peptidoglycan requires binding of DAP-PG by PGRP1 and the MBP N-terminal domain and association of the LDLa2–5 region of proHP14 with the MBP C-terminal domain. These interactions may concentrate the proHP14 zymogen at the bacterial cell wall surface and promote autoactivation.

Okra mucilage powder: a novel functional ingredient with antioxidant activity and antibacterial mode of action revealed by scanning and transmission electron microscopy

SummaryOkra mucilage powder (OMP) was firstly reported its antibacterial mode of action observed by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The reconstituted OMP or okra mucilage solution (OMS) with the concentration at 120 mg/mL was analysed for its total phenolic contents, total flavonoid contents and antioxidant activity reported as 11.33 ± 0.265 mg gallic acid/100 mL, 5.06 ± 0.197 mg catechin/100 mL and 176.18 ± 12.180 µmol Trolox equivalent/100 mL, respectively. The OMS exhibited strong antibacterial activities on Bacillus cereus, Staphylococcus aureus and Listeria monocytogenes with more than 3 log reduction from bacterial inoculum. Nonetheless, mode of action of OMS studied via TEM revealed that cells treated with OMS showed broken cell walls, cell membrane separation and intracellular content leakage. SEM photography revealed furrowed bacterial cell wall surface for this OMS‐treated cells when compared with untreated cells. Overall, our data suggested that the reconstituted OMP offered the greatest functional properties as antioxidant properties and antibacterial activities, aiding to its future application as a potential novel functional ingredient with health‐promoting application.

Astilbin Inhibits the Activity of Sortase A from Streptococcus mutans.

Streptococcus mutans (S. mutans) is the primary etiological agent of dental caries. The S. mutans enzyme sortase A (SrtA) is responsible for anchoring bacterial cell wall surface proteins involved in host cell attachment and biofilm formation. Thus, SrtA is an attractive target for inhibiting dental caries caused by S. mutans-associated acid fermentation. In this study, we observed that astilbin, a flavanone compound extracted from Rhizoma Smilacis Glabrae, has potent inhibitory activity against the S. mutans SrtA, with an IC50 of 7.5 μg/mL. In addition, astilbin was proven to reduce the formation of biofilm while without affecting the growth of S. mutans. The results of a molecular dynamics simulation and a mutation analysis revealed that the Arg213, Leu111, and Leu116 of SrtA are important for the interaction between SrtA and astilbin. The results of this study demonstrate the potential of using astilbin as a nonbactericidal agent to modulate pathogenicity of S. mutans by inhibiting the activity of SrtA.

Rhamnolipid-enhanced Pseudomonas putida biofilm formation on hydrophilic surfaces with toluene as the bacterium's sole carbon source

The effect of surfactants on biofilm formation depends on the surfactant type, the bacterial strain and the carrier. In order to enhance the bioremediation process in the presence of a biosurfactant, it is crucial to choose the appropriate bacterial strain and carrier for biofilm formation. In the current research, a linear correlation was observed between toluene degradation by a planktonic Pseudomonas putida culture and increasing rhamnolipid concentrations. Biofilm viability on the hydrophilic surface after 24 h was 0.3 ± 0.007 (OD 540 nm), 1.4 fold higher than on the hydrophobic surface. The hydrophobicity of the bacterial cell wall for cultures that were grown on toluene in the presence and in the absence of rhamnolipid was determined by the contact angle of water measurement on the bacterial layer as well as by the bacterial adherence to a solution of hydrocarbon (BATH test). The results showed that the bacterial cell wall surface in the presence and absence of rhamnolipid is highly hydrophilic. The viability of the biofilm which was grown on hydrophilic surfaces: treated ELISA plate and coal bottom ash was 7-fold and 13-fold higher, respectively, when the cultures were grown on toluene in the presence of rhamnolipid as compared with its absence. In conclusion, the combination of hydrophilic surfaces, rhamnolipid and P. putida may serve as a good choice for enhancing the bioremediation process of toluene.

Antibacterial effect of ultrafine nanodiamond against gram-negative bacteria Escherichia coli.

We investigate the antibacterial effect of ultrafine nanodiamond particles with an average size of 5 nm against the gram-negative bacteria Escherichia coli (E. coli). UV-visible, Raman spectroscopy, and scanning electron microscopy (SEM) have been employed to elucidate the nature of the interaction. The influence on bacterial growth was monitored by measuring optical densities of E. coli at 600 nm as a function of time in the presence of carboxylated nanodiamond (cND) particles (100 μg/ml ) in highly nutritious liquid Luria-Bertani medium. The SEM images prove that cND particles are attached to the bacterial cell wall surface and some portion of the bacterial cell wall undergoes destruction. Due to the change of the protein structure on the bacterial wall, a small Raman shift in the region of 1400 to 1700 cm⁻¹ was observed when E. coli interacted with cNDs. Raman mapping images show strong evidence of cND attachment at the bacterial cell wall surface. Electrotransformation of E. coli with a fluorescent protein markers experiment demonstrated that the interaction mechanisms are different for E. coli treated with cND particles, E. coli by lysozyme treatment, and E. coli that suffer lysis.

Molecular Basis of Ligand Recognition by Mammalian Peptidoglycan Recognition Protein

Short peptidoglycan recognition protein (PGRP-S) is a member of the innate immune system which provides the first line of defense to hosts against invading microbes. PGRP-S recognizes conserved motifs called pathogen associated molecular patterns (PAMPs) present in microorganisms but absent in host. We have determined the structure of camel peptidoglycan recognition protein (CPGRP-S) with various PAMPs like peptidoglycans, lipopolysaccharide, lipoteichoic acid, mycolic acid and five different fatty acids. The native structure revealed the presence of four crystallographically independent molecules A, B, C and D in the asymmetric unit. The buried surface area calculations indicated two stable contact regions, A-B and C-D which corresponded to opposite faces of the protein molecule resulting in the formation of a linear chain with alternating A-B and C-D contacts. This leads to the formation of multiple subsites contributed by molecules A, C, and D and a supporting diffusion channel formed by B and D molecules. The co-complexed structures indicated that the glycan moieties which contain PAMPs bind at C-D interface (PAMP-binding site) whereas the fatty acids occupy the cleft formed at A-B interface demonstrating that the binding sites for glycans and fatty acids are independent of each other. Similarly, the binary complex with the muramyl dipeptide and ternary complex with N-acetyl-D-glucosamine and beta-maltose revealed that these are accommodated in different sub-regions of the PAMP-binding site. This indicates that the PAMP-binding site is capable of accepting different kinds of PAMPs exhibiting varying specificities. It thus appears that the mode of binding of CPGRP-S essentially involves the interactions with the bacterial cell wall surface molecular patterns rather than cell membranes leading to the sequestration of the bacteria. Hence, CPGRP-S may be a protein antibiotic which may not suffer from bacterial resistance.

Delayed hyperacute rejection in a patient who developed clostridium difficile infection after ABO-incompatible kidney transplantation

Over the past decade ABO incompatible transplantation has emerged as an important potential source for increasing living kidney transplantation in selected transplant centers. Early reports suggest that patients who have elevated serum anti-blood group antibody titers (anti-A/B) before transplantation and a rebound antibody production after antibody removal may be at high immunological risk. With currently available immune modulation protocols and immunosuppressive therapy, excellent short- and long-term patient and graft survival rates have been achieved even in those with high anti-A/B antibody titers before plasmapheresis or immunoadsorption. Nonetheless, acute infection with an organism possessing surface markers analogous to blood group antigens such as carbohydrate structures on the surface of bacterial cell wall occurring before the firm establishment of accommodation can trigger the onset of acute antibody-mediated rejection. We herein report a case of delayed hyperacute rejection in an A1 to O, ABO incompatible transplant recipient following an episode of Clostridium difficile infection.

Self-Assembly of S-Layer-Enveloped Cytochrome c Polyelectrolyte Multilayers

We report a study of the electrostatic layer-by-layer self-assembly of electroactive polyelectrolyte multilayers incorporating the redox protein cytochrome c (cyt c) combined with recrystallization of the bacterial cell wall surface layer from Bacillus sphaericus CCM 2177 SbpA (S-layer). The polyelectrolyte multilayer assembly was prepared on flat gold electrodes with a nanometer-scale roughness that allowed monitoring of the film formation throughout all the assembly stages by atomic force microscopy measurements in liquid with respect to topography and forces. The deposition of alternating layers of sulfonated polyaniline and cyt c was carried out by adsorption from the corresponding solutions on a cyt c monolayer electrode. The electroactivity of cyt c within the assembly was confirmed by cyclic voltammetry. We showed that the surface properties of the electrode terminating layer change after each adsorption step accordingly. We also found that S-layer recrystallization on the top of the multilayer film was feasible while electroactivity of cyt c within a polyelectrolyte matrix was partially maintained. This approach offers a new strategy to design a biocompatible and permselective outer envelope of a polyelectrolyte multilayer, promising sensor applications.

Hydrous ferric oxide precipitation in the presence of nonmetabolizing bacteria: Constraints on the mechanism of a biotic effect

We have used room temperature and cryogenic 57Fe Mössbauer spectroscopy, powder X-ray diffraction (pXRD), mineral magnetometry, and transmission electron microscopy (TEM), to study the synthetic precipitation of hydrous ferric oxides (HFOs) prepared either in the absence (abiotic, a-HFO) or presence (biotic, b-HFO) of nonmetabolizing bacterial cells ( Bacillus subtilis or Bacillus licheniformis, ∼10 8 cells/mL) and under otherwise identical chemical conditions, starting from Fe(II) (10 −2, 10 −3, or 10 −4 mol/L) under open oxic conditions and at different pH (6–9). We have also performed the first Mössbauer spectroscopy measurements of bacterial cell wall ( Bacillus subtilis) surface complexed Fe, where Fe(III) (10 −3.5–10 −4.5 mol/L) was added to a fixed concentration of cells (∼10 8 cells/mL) under open oxic conditions and at various pH (2.5–4.3). We find that non-metabolic bacterial cell wall surface complexation of Fe is not passive in that it affects Fe speciation in at least two ways: (1) it can reduce Fe(III) to sorbed-Fe 2+ by a proposed steric and charge transfer effect and (2) it stabilizes Fe(II) as sorbed-Fe 2+ against ambient oxidation. The cell wall sorption of Fe occurs in a manner that is not compatible with incorporation into the HFO structure (different coordination environment and stabilization of the ferrous state) and the cell wall-sorbed Fe is not chemically bonded to the HFO particle when they coexist (the sorbed Fe is not magnetically polarized by the HFO particle in its magnetically ordered state). This invalidates the concept that sorption is the first step in a heterogeneous nucleation of HFO onto bacterial cell walls. Both the a-HFOs and the b-HFOs are predominantly varieties of ferrihydrite (Fh), often containing admixtures of nanophase lepidocrocite (nLp), yet they show significant abiotic/biotic differences: Biotic Fh has less intraparticle (including surface region) atomic order (Mössbauer quadrupole splitting), smaller primary particle size (magnetometry blocking temperature), weaker Fe to particle bond strength (Mössbauer center shift), and no six-line Fh (6L-Fh) admixture (pXRD, magnetometry). Contrary to current belief, we find that 6L-Fh appears to be precipitated directly, under a-HFO conditions, from either Fe(II) or Fe(III), and depending on Fe concentration and pH, whereas the presence of bacteria disables all such 6L-Fh precipitation and produces two-line Fh (2L-Fh)-like biotic coprecipitates. Given the nature of the differences between a-HFO and b-HFO and their synthesis condition dependences, several biotic precipitation mechanisms (template effect, near-cell environment effect, catalyzed nucleation and/or growth effect, and substrate-based coprecipitation) are ruled out. The prevailing present view of a template or heterogeneous nucleation barrier reduction effect, in particular, is shown not to be the cause of the large observed biotic effects on the resulting HFOs. The only proposed mechanism (relevant to Fh) that is consistent with all our observations is coprecipitation with and possible surface poisoning by ancillary bacteriagenic compounds. That bacterial cell wall functional groups are redox active and the characteristics of biotic (i.e., natural) HFOs compared to those of abiotic (i.e., synthetic) HFOs have several possible biogeochemical implications regarding Fe cycling, in the photic zones of water columns in particular.