Analysis of Interactions Occurring during the Pyrolysis of Lignocellulosic Biomass.

This paper presents a review of the recent advances in research on the interactions between the components of lignocellulosic biomass. The literature reports on the effects of interaction between lignocellulosic biomass components, such as cellulose-lignin, lignin-hemicellulose, and hemicellulose-cellulose, were discussed. The results obtained by other researchers were analyzed from the viewpoint of the interactions between the pyrolysis products formed along with the impact effects of the organic and inorganic components present or added to the biomass with regard to the yield and composition of the pyrolysis products. Disagreements about some statements were noted along with the lack of an unequivocal opinion about the directivity of interactions occurring during biomass pyrolysis. Based on the data in the scientific literature, it was suggested that the course of the pyrolysis process of biomass blends can be appropriately directed by changes in the ratio of basic biomass components or by additions of inorganic or organic substances.

Prevotella intermedia produces two proteins homologous to Porphyromonas gingivalis HmuY but with different heme coordination mode.

As part of the infective process, Porphyromonas gingivalis must acquire heme which is indispensable for life and enables the microorganism to survive and multiply at the infection site. This oral pathogenic bacterium uses a newly discovered novel hmu heme uptake system with a leading role played by the HmuY hemophore-like protein, responsible for acquiring heme and increasing virulence of this periodontopathogen. We demonstrated that Prevotella intermedia produces two HmuY homologs, termed PinO and PinA. Both proteins were produced at higher mRNA and protein levels when the bacterium grew under low-iron/heme conditions. PinO and PinA bound heme, but preferentially under reducing conditions, and in a manner different from that of the P. gingivalis HmuY. The analysis of the three-dimensional structures confirmed differences between apo-PinO and apo-HmuY, mainly in the fold forming the heme-binding pocket. Instead of two histidine residues coordinating heme iron in P. gingivalis HmuY, PinO and PinA could use one methionine residue to fulfill this function, with potential support of additional methionine residue/s. The P. intermedia proteins sequestered heme only from the host albumin-heme complex under reducing conditions. Our findings suggest that HmuY-like family might comprise proteins subjected during evolution to significant diversification, resulting in different heme coordination modes. The newer data presented in this manuscript on HmuY homologs produced by P. intermedia sheds more light on the novel mechanism of heme uptake, could be helpful in discovering their biological function, and in developing novel therapeutic approaches.

Antimicrobial activity of stable hemiaminals against Porphyromonas gingivalis.

Porphyromonas gingivalis is a major etiologic agent and a key pathogen responsible for the development and progression of chronic periodontitis. Controlling the number of periodontal pathogens is one of the primary actions for maintaining oral health; therefore, active compounds with a capacity to exert antimicrobial activity have received considerable attention as they may represent potential new therapeutic agents for the treatment of chronic periodontitis. Heterocyclic compounds possessing 1,2,4- or 1,2,3-triazoles are known for several biological activities, including antibacterial properties. Among them are stable hemiaminals which can be obtained in reaction between nitrobenzaldehyde derivatives and 4-amino-1,2,4-triazole or 4-amino-3,5-dimethyl-1,2,4-triazole. In this study, we selected two relatively stable hemiaminals: (2,4-dinitrophenyl)(4H-1,2,4-triazole-4-ylamino)methanol (24DNTAM) and (2,4-dinitrophenyl)(4H-3,5-dimethyl-1,2,4-triazole-4-ylamino)methanol (24DNDMTAM). Both compounds showed promising anti-P. gingivalis activity, higher against ATCC 33277 strain as compared to A7436 strain. The lowest hemiaminal concentration inhibiting visible planktonic bacterial growth under high-iron/heme conditions was ∼0.06 mg/ml, and the lowest hemiaminal concentration showing killing of bacteria was ∼0.25 mg/ml. Antimicrobial activity was also observed against P. gingivalis grown on blood agar plates. Slightly higher antimicrobial activity of both compounds was observed when P. gingivalis was grown in co-cultures with epithelial HeLa cells under low-iron/heme conditions, which mimic those occurring in vivo. 24DNTAM was more effective against P. gingivalis, but exhibited higher cytotoxic activity against epithelial and red blood cells, as compared with 24DNDMTAM. We conclude that both hemiaminals might originate a novel group of biologically important molecules.

Differential roles of tryptophan residues in conformational stability of Porphyromonas gingivalis HmuY hemophore.

BACKGROUND: We have previously shown that the P. gingivalis HmuY hemophore-like protein binds heme and scavenges heme from host hemoproteins to further deliver it to the cognate heme receptor HmuR. The aim of this study was to characterize structural features of HmuY variants in the presence and absence of heme with respect to roles of tryptophan residues in conformational stability. RESULTS: HmuY possesses tryptophan residues at positions 51 and 73, which are conserved in HmuY homologs present in a variety of bacteria, and a tryptophan residue at position 161, which has been found only in HmuY identified in P. gingivalis strains. We expressed and purified the wildtype HmuY and its protein variants with single tryptophan residues replaced by alanine or tyrosine residues. All HmuY variants were subjected to thermal denaturation and fluorescence spectroscopy analyses. Replacement of the most buried W161 only moderately affects protein stability. The most profound effect of the lack of a large hydrophobic side chain in respect to thermal stability is observed for W73. Also replacement of the W51 exposed on the surface results in the greatest loss of protein stability and even the large aromatic side chain of a tyrosine residue has little potential to substitute this tryptophan residue. Heme binding leads to different exposure of the tryptophan residue at position 51 to the surface of the protein. Differences in structural stability of HmuY variants suggest the change of the tertiary structure of the protein upon heme binding. CONCLUSIONS: Here we demonstrate differential roles of tryptophan residues in the protein conformational stability. We also propose different conformations of apo- and holoHmuY caused by tertiary changes which allow heme binding to the protein.

Preliminary assessment of the psychometric properties of the Polish version of the Questionnaire to Assess Affective and Cognitive Empathy (QAACE) in Children.

BACKGROUND: This article reports the Polish adaptation of the Questionnaire to Assess Affective and Cognitive Empathy (QAACE) by Zoll and Enz - a multidimensional self-report questionnaire used to measure empathy in children aged 8-14. The QAACE is based on a two-factor cognitive-emotional model of empathy. It has a number of international adaptations and offers a convenient Polish-language tool for use with young children and adolescents. PARTICIPANTS AND PROCEDURE: The sample consisted of 677 children aged 8-13. The survey was conducted on school premises, during classes, by an appropriately prepared researcher. RESULTS: Confirmatory factor analysis revealed a good fitting measurement model representing the original underlying factor structure of the QAACE among Polish children. The reliability of the questionnaire as measured by Cronbach's α and McDonald's ω was good. The reliability of the scale as assessed by the test-retest method (after four weeks) was .80. We assessed the validity of the tool by analyzing the correlation of empathy with love and sadism. General empathy, as well as cognitive and affective empathy, is positively related to love. The hypothesis that sadism is significantly related to empathy was also partially confirmed. General empathy and affective empathy are negatively correlated with sadism, while there was no relationship between sadism and cognitive empathy. CONCLUSIONS: The questionnaire is the first widely available tool of this type to examine empathy and its components appropriate for children and adolescents in Poland. The questionnaire can be a useful screening test for detecting children's level of empathy.

A Comparative Study on Laser Powder Bed Fusion of Differently Atomized 316L Stainless Steel.

The significant growth of Additive Manufacturing (AM), visible over the last ten years, has driven an increase in demand for small gradation metallic powders of a size lower than 100 µm. Until now, most affordable powders for AM have been produced using gas atomization. Recently, a new, alternative method of powder production based on ultrasonic atomization with melting by electric arc has appeared. This paper summarizes the preliminary research results of AM samples made of two AISI 316L steel powder batches, one of which was obtained during Ultrasonic Atomization (UA) and the other during Plasma Arc Gas Atomization (PAGA). The comparison starts from powder particle statistical distribution, chemical composition analysis, density, and flowability measurements. After powder analysis, test samples were produced using AM to observe the differences in microstructure, porosity, and hardness. Finally, the test campaign covered an analysis of mechanical properties, including tensile testing with Digital Image Correlation (DIC) and Charpy's impact tests. A comparative study of parts made of ultrasonic and gas atomization powders confirms the likelihood that both methods can deliver material of similar properties.

Iron(III) mesoporphyrin IX and iron(III) deuteroporphyrin IX bind to the Porphyromonas gingivalis HmuY hemophore.

Porphyromonas gingivalis acquires heme through an outer-membrane heme transporter HmuR and heme-binding hemophore-like lipoprotein HmuY. Here, we compare binding of iron(III) mesoporphyrin IX (mesoheme) and iron(III) deuteroporphyrin IX (deuteroheme) to HmuY with that of iron(III) protoporphyrin IX (protoheme) and protoporphyrin IX (PPIX) using spectroscopic methods. In contrast to PPIX, mesoheme and deuteroheme enter the HmuY heme cavity and are coordinated by His134 and His166 residues in a fully analogous way to protoheme binding. However, in the case of deuteroheme two forms of HmuY-iron porphyrin complex were observed differing by a 180° rotation of porphyrin about the α-γ-meso-carbon axis. Since the use of porphyrins either as active photosensitizers or in combination with antibiotics may have therapeutic value for controlling bacterial growth in vivo, it is important to compare the binding of heme derivatives to HmuY.

Sensors to Detect Sarin Simulant.

This work presents a literature review concerning the construction, properties and application of different sensors used to detect dimethyl methylphonate (DMMP), which is the simulant of sarin. Sensors sensitive to mass are described, together with such sensors as: SAW, QCM, MEMS, also chemical capacitors, semiconductors, and field effect transistors.

Porphyromonas gingivalis PgFur Is a Member of a Novel Fur Subfamily With Non-canonical Function.

Porphyromonas gingivalis, a keystone pathogen of chronic periodontitis, uses ferric uptake regulator homolog (PgFur) to regulate production of virulence factors. This study aimed to characterize PgFur protein in regard to its structure-function relationship. We experimentally identified the 5' mRNA sequence encoding the 171-amino-acid-long PgFur protein in the A7436 strain and examined this PgFur version as a full-length protein. PgFur protein did not bind to the canonical Escherichia coli Fur box, but the wild-type phenotype of the mutant Δpgfur strain was restored partially when expression of the ecfur gene was induced from the native pgfur promoter. The full-length PgFur protein contained one zinc atom per protein monomer, but did not bind iron, manganese, or heme. Single cysteine substitutions of CXXC motifs resulted in phenotypes similar to the mutant Δpgfur strain. The modified proteins were produced in E. coli at significantly lower levels, were highly unstable, and did not bind zinc. The pgfur gene was expressed at the highest levels in bacteria cultured for 24 h in the absence of iron and heme or at higher levels in bacteria cultured for 10 h in the presence of protoporphyrin IX source. No influence of high availability of Fe2+, Zn2+, or Mn2+ on pgfur gene expression was observed. Two chromosomal mutant strains producing protein lacking 4 (pgfurΔ4aa) or 13 (pgfurΔ13aa) C-terminal amino acid residues were examined in regard to importance of the C-terminal lysine-rich region. The pgfurΔ13aa strain showed a phenotype typical for the mutant Δpgfur strain, but both the wild-type PgFur protein and its truncated version bound zinc with similar ability. The Δpgfur mutant strain produced higher amounts of HmuY protein compared with the wild-type strain, suggesting compromised regulation of its expression. Potential PgFur ligands, Fe2+, Mn2+, Zn2+, PPIX, or serum components, did not influence HmuY production in the Δpgfur mutant strain. The mutant pgfurΔ4aa and pgfurΔ13aa strains exhibited affected HmuY protein production. PgFur, regardless of the presence of the C-terminal lysine-rich region, bound to the hmu operon promoter. Our data suggest that cooperation of PgFur with partners/cofactors and/or protein/DNA modifications would be required to accomplish its role played in an in vivo multilayer regulatory network.

Tannerella forsythia Tfo belongs to Porphyromonas gingivalis HmuY-like family of proteins but differs in heme-binding properties.

Porphyromonas gingivalis is considered the principal etiologic agent and keystone pathogen of chronic periodontitis. As an auxotrophic bacterium, it must acquire heme to survive and multiply at the infection site. P. gingivalis HmuY is the first member of a novel family of hemophore-like proteins. Bacterial heme-binding proteins usually use histidine-methionine or histidine-tyrosine residues to ligate heme-iron, whereas P. gingivalis HmuY uses two histidine residues. We hypothesized that other 'red complex' members, i.e. Tannerella forsythia and Treponema denticola might utilize similar heme uptake mechanisms to the P. gingivalis HmuY. Comparative and phylogenetic analyses suggested differentiation of HmuY homologs and low conservation of heme-coordinating histidine residues present in HmuY. The homologs were subjected to duplication before divergence of Bacteroidetes lineages, which could facilitate evolution of functional diversification. We found that T. denticola does not code an HmuY homolog. T. forsythia protein, termed as Tfo, binds heme, but preferentially in the ferrous form, and sequesters heme from the albumin-heme complex under reducing conditions. In agreement with that, the 3D structure of Tfo differs from that of HmuY in the folding of heme-binding pocket, containing two methionine residues instead of two histidine residues coordinating heme in HmuY. Heme binding to apo-HmuY is accompanied by movement of the loop carrying the His166 residue, closing the heme-binding pocket. Molecular dynamics simulations (MD) demonstrated that this conformational change also occurs in Tfo. In conclusion, our findings suggest that HmuY-like family might comprise proteins subjected during evolution to significant diversification, resulting in different heme-binding properties.

Anti-HmuY antibodies specifically recognize Porphyromonas gingivalis HmuY protein but not homologous proteins in other periodontopathogens.

Given the emerging evidence of an association between periodontal infections and systemic conditions, the search for specific methods to detect the presence of P. gingivalis, a principal etiologic agent in chronic periodontitis, is of high importance. The aim of this study was to characterize antibodies raised against purified P. gingivalis HmuY protein and selected epitopes of the HmuY molecule. Since other periodontopathogens produce homologs of HmuY, we also aimed to characterize responses of antibodies raised against the HmuY protein or its epitopes to the closest homologous proteins from Prevotella intermedia and Tannerella forsythia. Rabbits were immunized with purified HmuY protein or three synthetic, KLH-conjugated peptides, derived from the P. gingivalis HmuY protein. The reactivity of anti-HmuY antibodies with purified proteins or bacteria was determined using Western blotting and ELISA assay. First, we found homologs of P. gingivalis HmuY in P. intermedia (PinO and PinA proteins) and T. forsythia (Tfo protein) and identified corrected nucleotide and amino acid sequences of Tfo. All proteins were overexpressed in E. coli and purified using ion-exchange chromatography, hydrophobic chromatography and gel filtration. We demonstrated that antibodies raised against P. gingivalis HmuY are highly specific to purified HmuY protein and HmuY attached to P. gingivalis cells. No reactivity between P. intermedia and T. forsythia or between purified HmuY homologs from these bacteria and anti-HmuY antibodies was detected. The results obtained in this study demonstrate that P. gingivalis HmuY protein may serve as an antigen for specific determination of serum antibodies raised against this bacterium.

The Porphyromonas gingivalis HmuY haemophore binds gallium(iii), zinc(ii), cobalt(iii), manganese(iii), nickel(ii), and copper(ii) protoporphyrin IX but in a manner different to iron(iii) protoporphyrin IX.

Porphyromonas gingivalis, a major etiological agent of chronic periodontitis, acquires haem from host haemoproteins through a haem transporter HmuR and a haemophore HmuY. The aim of this study was to analyse the binding specificity of HmuY towards non-iron metalloporphyrins which may be employed as antimicrobials to treat periodontitis. HmuY binds gallium(iii), zinc(ii), cobalt(iii), manganese(iii), nickel(ii), and copper(ii) protoporphyrin IX but in a manner different to iron(iii) protoporphyrin IX which uses His(134) and His(166) as axial ligands. The metal ions in Ga(iii)PPIX and Zn(ii)PPIX can accept only His(166) as an axial ligand, whereas nickel(ii) and copper(ii) interact exclusively with His(134). Two forms of pentacoordinate manganese(iii) are present in the Mn(iii)PPIX-HmuY complex since the metal accepts either His(134) or His(166) as a single axial ligand. The cobalt ion is hexacoordinate in the Co(iii)PPIX-HmuY complex and binds His(134) and His(166) as axial ligands; however, some differences in their environments exist. Despite different coordination modes of the central metal ion, gallium(iii), zinc(ii), cobalt(iii), and manganese(iii) protoporphyrin IX bound to the HmuY haemophore cannot be displaced by excess haem. All of the metalloporphyrins examined bind to a P. gingivalis wild-type strain with higher ability compared to a mutant strain lacking a functional hmuY gene, thus corroborating binding of non-iron metalloporphyrins to purified HmuY protein. Our results further clarify the basis of metalloporphyrin acquisition by P. gingivalis and add to understanding of the interactions with porphyrin derivatives which exhibit antimicrobial activity against P. gingivalis.

The unique hmuY gene sequence as a specific marker of Porphyromonas gingivalis.

Porphyromonas gingivalis, a major etiological agent of chronic periodontitis, acquires heme from host hemoproteins using the HmuY hemophore. The aim of this study was to develop a specific P. gingivalis marker based on a hmuY gene sequence. Subgingival samples were collected from 66 patients with chronic periodontitis and 40 healthy subjects and the entire hmuY gene was analyzed in positive samples. Phylogenetic analyses demonstrated that both the amino acid sequence of the HmuY protein and the nucleotide sequence of the hmuY gene are unique among P. gingivalis strains/isolates and show low identity to sequences found in other species (below 50 and 56%, respectively). In agreement with these findings, a set of hmuY gene-based primers and standard/real-time PCR with SYBR Green chemistry allowed us to specifically detect P. gingivalis in patients with chronic periodontitis (77.3%) and healthy subjects (20%), the latter possessing lower number of P. gingivalis cells and total bacterial cells. Isolates from healthy subjects possess the hmuY gene-based nucleotide sequence pattern occurring in W83/W50/A7436 (n = 4), 381/ATCC 33277 (n = 3) or TDC60 (n = 1) strains, whereas those from patients typically have TDC60 (n = 21), W83/W50/A7436 (n = 17) and 381/ATCC 33277 (n = 13) strains. We observed a significant correlation between periodontal index of risk of infectiousness (PIRI) and the presence/absence of P. gingivalis (regardless of the hmuY gene-based sequence pattern of the isolate identified [r = 0.43; P = 0.0002] and considering particular isolate pattern [r = 0.38; P = 0.0012]). In conclusion, we demonstrated that the hmuY gene sequence or its fragments may be used as one of the molecular markers of P. gingivalis.