Dong-Heon Baek

A new classification system for bacterial Rieske non-heme iron aromatic ring-hydroxylating oxygenases

BackgroundRieske non-heme iron aromatic ring-hydroxylating oxygenases (RHOs) are multi-component enzyme systems that are remarkably diverse in bacteria isolated from diverse habitats. Since the first classification in 1990, there has been a need to devise a new classification scheme for these enzymes because many RHOs have been discovered, which do not belong to any group in the previous classification. Here, we present a scheme for classification of RHOs reflecting new sequence information and interactions between RHO enzyme components.ResultWe have analyzed a total of 130 RHO enzymes in which 25 well-characterized RHO enzymes were used as standards to test our hypothesis for the proposed classification system. From the sequence analysis of electron transport chain (ETC) components of the standard RHOs, we extracted classification keys that reflect not only the phylogenetic affiliation within each component but also relationship among components. Oxygenase components of standard RHOs were phylogenetically classified into 10 groups with the classification keys derived from ETC components. This phylogenetic classification scheme was converted to a new systematic classification consisting of 5 distinct types. The new classification system was statistically examined to justify its stability. Type I represents two-component RHO systems that consist of an oxygenase and an FNRC-type reductase. Type II contains other two-component RHO systems that consist of an oxygenase and an FNRN-type reductase. Type III represents a group of three-component RHO systems that consist of an oxygenase, a [2Fe-2S]-type ferredoxin and an FNRN-type reductase. Type IV represents another three-component systems that consist of oxygenase, [2Fe-2S]-type ferredoxin and GR-type reductase. Type V represents another different three-component systems that consist of an oxygenase, a [3Fe-4S]-type ferredoxin and a GR-type reductase.ConclusionThe new classification system provides the following features. First, the new classification system analyzes RHO enzymes as a whole. RwithSecond, the new classification system is not static but responds dynamically to the growing pool of RHO enzymes. Third, our classification can be applied reliably to the classification of incomplete RHOs. Fourth, the classification has direct applicability to experimental work. Fifth, the system provides new insights into the evolution of RHO systems based on enzyme interaction.

Polycyclic Aromatic Hydrocarbon Metabolic Network in Mycobacterium vanbaaleniiPYR-1

This study investigated a metabolic network (MN) from Mycobacterium vanbaalenii PYR-1 for polycyclic aromatic hydrocarbons (PAHs) from the perspective of structure, behavior, and evolution, in which multilayer omics data are integrated. Initially, we utilized a high-throughput proteomic analysis to assess the protein expression response of M. vanbaalenii PYR-1 to seven different aromatic compounds. A total of 3,431 proteins (57.38% of the genome-predicted proteins) were identified, which included 160 proteins that seemed to be involved in the degradation of aromatic hydrocarbons. Based on the proteomic data and the previous metabolic, biochemical, physiological, and genomic information, we reconstructed an experiment-based system-level PAH-MN. The structure of PAH-MN, with 183 metabolic compounds and 224 chemical reactions, has a typical scale-free nature. The behavior and evolution of the PAH-MN reveals a hierarchical modularity with funnel effects in structure/function and intimate association with evolutionary modules of the functional modules, which are the ring cleavage process (RCP), side chain process (SCP), and central aromatic process (CAP). The 189 commonly upregulated proteins in all aromatic hydrocarbon treatments provide insights into the global adaptation to facilitate the PAH metabolism. Taken together, the findings of our study provide the hierarchical viewpoint from genes/proteins/metabolites to the network via functional modules of the PAH-MN equipped with the engineering-driven approaches of modularization and rationalization, which may expand our understanding of the metabolic potential of M. vanbaalenii PYR-1 for bioremediation applications.

Restoration of peri-implant defects in immediate implant installations by Choukroun platelet-rich fibrin and silk fibroin powder combination graft.

OBJECTIVE The objective of this study was to determine the capability of silk fibroin powder as a biomaterial template for the restoration of peri-implant defects when mixed with Choukroun platelet-rich fibrin (PRF) in vivo. STUDY DESIGN Ten New Zealand white rabbits were used for this study. Using a trephine bur (diameter 7.0 mm), 2 monocortical defects were prepared. Subsequently, 2 dental implants were installed into the tibia (diameter 3.0 mm, length 10.0 mm). In the experimental group, the peri-implant defect was filled with a combination graft of silk fibroin powder and Choukroun PRF. The control was left in an unfilled state. The animals were killed at 8 weeks. Subsequently, a removal torque test and a histomorphometric analysis were done. RESULTS The removal torque for the experimental group was 30.34 +/- 5.06 N.cm, whereas it was 21.86 +/- 3.39 N.cm for the control. The difference between the 2 groups was statistically significant (P = .010). Mean new bone formation was 51.93 +/- 27.90% in the experimental group and 11.67 +/- 15.12% in the control (P = .003). Mean bone-to-implant contact was 43.07 +/- 21.96% in the experimental group and 15.37 +/- 23.84% in the control (P = .002). CONCLUSION A peri-implant defect can be successfully repaired by the application of Choukroun PRF and silk fibroin powder.

Antibacterial and Neutralizing Effect of Human β-Defensins on Enterococcus faecalis and Enterococcus faecalis Lipoteichoic Acid

INTRODUCTION Enterococcus faecalis is frequently found in the root canal of teeth, is a major microorganism of endodontic therapy failure, and is associated with chronic apical periodontitis. Human β-defensins (HBDs) are known to play critical roles in defending the host against infectious microbes and producing dental pulp in healthy and patients. The purpose of the present study was to investigate the bactericidal and neutralizing effects of HBDs on E. faecalis and E. faecalis lipoteichoic acid (Ef LTA) as a major virulence factor of E. faecalis. METHODS HBD-1, -2, -3, and -4 were synthesized and investigated the susceptibility against E. faecalis. Also, the neutralizing effects of HBDs on cytokine and intercellular adhesion molecule 1 (ICAM-1) expression by activity of E. faecalis and Ef LTA were analyzed using enzyme-linked immunosorbent assay and flow cytometry. RESULTS HBD-1 and -2 were weakly susceptible, and HBD-3 and HBD-4 were strongly susceptible to E. faecalis. All of the HBDs exhibited neutralizing effects on the activity of Ef LTA, and HBD-3 strongly neutralized the activity of E. faecalis in tumor necrosis factor-α, interleukin-8, and ICAM-1 expression. The neutralizing effects of HBDs were to inhibit E. faecalis or Ef LTA binding to the host cells. CONCLUSIONS These results suggest that the induction of HBDs might have great potential as endodontic therapeutic agents.

Aerosol Deposition of Hydroxyapatite and 4-Hexylresorcinol Coatings on Titanium Alloys for Dental Implants

PURPOSE Aerosol deposition is a newly developed technique, and it can deliver the drug from a hydroxyapatite (HA)-coated surface. 4-Hexylresorcinol (4-HR) is a well-known antiseptic. The influence of the 4-HR component of HA coatings on titanium surfaces was studied in vitro and in vivo. MATERIALS AND METHODS X-ray diffraction and Fourier transform infrared techniques were used for the evaluation of the coating. The cellular response of the coating was evaluated by scanning electron microscopic study, MTT assay, alkaline phosphatase assay, and osteocalcin assay. In addition, the dental implant was coated with HA or HA + 4-HR. The implant was installed into the tibia of a rabbit after contamination by Aggregatibacter actinomycetemcomitans. The torque test and histologic analysis were then performed at 8 weeks after the operation. RESULTS By use of an aerosol deposition technique, the combination of HA and 4-HR was successfully coated onto a titanium surface, which was confirmed by x-ray diffraction and Fourier transform infrared techniques. MG63 cells attached more rapidly to the HA + 4-HR coating than to the HA-only coating. The HA + 4-HR coating had significantly increased osteocalcin expression and alkaline phosphatase activity compared with the HA-only coating (P < .05). The dental implant coated with HA + 4-HR had a significantly higher removal torque value than that coated with HA alone at 8 weeks after surgery (P < .05). On histologic analysis, both the bone formation value and the bone-to-implant contact value were significantly higher in the HA + 4-HR group than in the HA-only group at 8 weeks after surgery (P < .05). CONCLUSIONS Collectively, the HA + 4-HR-coated dental implant had clear advantages over the HA-coated dental implant. Therefore HA + 4-HR coatings can be considered for patients who need immediate implant installation after tooth extraction or who have poor-quality bone.

Comparative functional pan-genome analyses to build connections between genomic dynamics and phenotypic evolution in polycyclic aromatic hydrocarbon metabolism in the genus Mycobacterium

BackgroundThe bacterial genus Mycobacterium is of great interest in the medical and biotechnological fields. Despite a flood of genome sequencing and functional genomics data, significant gaps in knowledge between genome and phenome seriously hinder efforts toward the treatment of mycobacterial diseases and practical biotechnological applications. In this study, we propose the use of systematic, comparative functional pan-genomic analysis to build connections between genomic dynamics and phenotypic evolution in polycyclic aromatic hydrocarbon (PAH) metabolism in the genus Mycobacterium.ResultsPhylogenetic, phenotypic, and genomic information for 27 completely genome-sequenced mycobacteria was systematically integrated to reconstruct a mycobacterial phenotype network (MPN) with a pan-genomic concept at a network level. In the MPN, mycobacterial phenotypes show typical scale-free relationships. PAH degradation is an isolated phenotype with the lowest connection degree, consistent with phylogenetic and environmental isolation of PAH degraders. A series of functional pan-genomic analyses provide conserved and unique types of genomic evidence for strong epistatic and pleiotropic impacts on evolutionary trajectories of the PAH-degrading phenotype. Under strong natural selection, the detailed gene gain/loss patterns from horizontal gene transfer (HGT)/deletion events hypothesize a plausible evolutionary path, an epistasis-based birth and pleiotropy-dependent death, for PAH metabolism in the genus Mycobacterium. This study generated a practical mycobacterial compendium of phenotypic and genomic changes, focusing on the PAH-degrading phenotype, with a pan-genomic perspective of the evolutionary events and the environmental challenges.ConclusionsOur findings suggest that when selection acts on PAH metabolism, only a small fraction of possible trajectories is likely to be observed, owing mainly to a combination of the ambiguous phenotypic effects of PAHs and the corresponding pleiotropy- and epistasis-dependent evolutionary adaptation. Evolutionary constraints on the selection of trajectories, like those seen in PAH-degrading phenotypes, are likely to apply to the evolution of other phenotypes in the genus Mycobacterium.

ClassRHO: a platform for classification of bacterial rieske non-heme iron ring-hydroxylating oxygenases.

We have developed an easy-to-use multiplatform classification tool, ClassRHO, which facilitates classification and comparison of bacterial Rieske non-heme iron aromatic ring-hydroxylating oxygenases (RHOs). Visualization and analysis can be generated on-the-fly by entering or uploading RHO query sequences. Pre-computed classifications were implemented for 42 standard RHO sequences. These 42 RHO sequences can be flexibly selected based on user requests. ClassRHO provides users with many options to view and analyze RHO sequences.

Effects of Streptococcus thermophilus on volatile sulfur compounds produced by Porphyromonas gingivalis

Halitosis as oral malodour is an unpleasant odour caused by volatile sulfur compounds (VSCs). VSCs are produced primarily by anaerobic bacteria that abundantly produce proteinase as trypsin-like enzyme. General therapies, such as mouthwash and plaque control, do not provide a continuous effect on oral halitosis. Streptococcus thermophilus is a probiotic bacterium that is beneficial for human health. The aim of this study was to evaluate the effect of S. thermophilus on Porphyromonas gingivalis-producing VSCs and to analyze the inhibitory mechanism of halitosis. P. gingivalis was cultured with or without S. thermophilus, and the emission of VSCs from the spent culture medium was measured by gas chromatography. In order to analyze the inhibitory effect, the antibacterial activity of S. thermophilus against P. gingivalis was assessed. After the spent culture medium or whole bacterial of S. thermophilus was mixed with the spent culture medium of P. gingivalis, VSCs were again measured by gas chromatograph. When S. thermophilus and P. gingivalis were co-cultivated, VSCs were present at a lower level than those of single-cultured P. gingivalis. S. thermophilus inhibited growth of P. gingivalis, and the whole bacteria and the spent culture medium of S. thermophilus reduced emission of VSCs gas. S. thermophilus may reduce oral malodour by inhibition of P. gingivalis growth and neutralizing VSCs with their metabolites or themselves.

Development of strain-specific PCR primers for the identification of Fusobacterium nucleatum subsp. fusiforme ATCC 51190T and subsp. vincentii ATCC 49256T

The objective of this study was to develop the strain-specific PCR primers for Fusobacterium nucleatum subsp. fusiforme ATCC 51190(T) and F. nucleatum subsp. vincentii ATCC 49256(T) based on the nucleotide sequence of the Fs17 and Fv35 DNA probes, respectively. The strain specificity was tested against 10 type strains of Fusobacterium spp. or subsp., 21 clinical isolates of F. nucleatum from Koreans, and five type strains of distinct Fusobacterium species. Primer sensitivity was determined by testing serial dilutions (4 ng-4 fg) of the purified genomic DNA from each of the type strains. PCR showed that two pairs of PCR primers, Fs17-F14/Fs17-R14 and Fv35-F1/Fv35-R1 primers, could produce strain-specific amplicons from F. nucleatum subsp. fusiforme ATCC 51190(T) and F. nucleatum subsp. vincentii ATCC 49256(T), respectively. The two PCR primer sets could detect as little as 0.4 pg or 4 pg of the genomic DNA of each target strain. These results suggest that the two sets of PCR primers could be used to identify F. nucleatum subsp. fusiforme ATCC 51190(T) and F. nucleatum subsp. vincentii ATCC 49256(T), particularly for ascertaining the authenticity of the strain.

Inhibitory effect of Lactococcus lactis on the bioactivity of periodontopathogens

Lactococcus lactis is a probiotic bacterium that produces various bacteriocins. Periodontopathogens induce inflammation and halitosis through the actions of lipopolysaccharide (LPS) and trypsin-like enzymes. The purpose of this study was to investigate the inhibitory effects of L. lactis on the bioactivity of periodontopathogens. To investigate the antimicrobial peptide of L. lactis, the spent culture medium (SCM) of L. lactis was treated with or without proteinase K after collection by centrifugation, and the antibacterial activity of SCM against periodontopathogens was assessed. To evaluate the neutralizing effect of L. lactis on halitosis, SCM of periodontopathogens was mixed with an L. lactis suspension, and the levels of volatile sulfur compounds (VSCs) were measured by gas chromatography. LPS from the periodontopathogens was extracted by an LPS extraction kit with little modification, and THP-1 cells as a monocytic cell line were treated with the extracted LPS in the presence or absence of UV-killed L. lactis. The production of inflammatory cytokines was analyzed by ELISA. The SCM of L. lactis exhibited antimicrobial activity against the periodontopathogens, whereas the proteinase K-treated SCM showed little antimicrobial activity. In addition, the L. lactis suspension had a neutralizing effect on the VSCs produced by periodontopathogens, and UV-killed L. lactis inhibited the production of IL-6 and TNF-α induced by the LPS. These results suggest that L. lactis may be a useful probiotic to prevent and treat periodontitis and halitosis.