Structural and sequence comparisons of bacterial enoyl-CoA isomerase and enoyl-CoA hydratase

Crystal structures of enoyl-coenzyme A (CoA) isomerase from Bosea sp. PAMC 26642 (BoECI) and enoyl-CoA hydratase from Hymenobacter sp. PAMC 26628 (HyECH) were determined at 2.35 and 2.70 Å resolution, respectively. BoECI and HyECH are members of the crotonase superfamily and are enzymes known to be involved in fatty acid degradation. Structurally, these enzymes are highly similar except for the orientation of their C-terminal helix domain. Analytical ultracentrifugation was performed to determine the oligomerization states of BoECI and HyECH revealing they exist as trimers in solution. However, their putative ligand-binding sites and active site residue compositions are dissimilar. Comparative sequence and structural analysis revealed that the active site of BoECI had one glutamate residue (Glu135), this site is occupied by an aspartate in some ECIs, and the active sites of HyECH had two highly conserved glutamate residues (Glu118 and Glu138). Moreover, HyECH possesses a salt bridge interaction between Glu98 and Arg152 near the active site. This interaction may allow the catalytic Glu118 residue to have a specific conformation for the ECH enzyme reaction. This salt bridge interaction is highly conserved in known bacterial ECH structures and ECI enzymes do not have this type of interaction. Collectively, our comparative sequential and structural studies have provided useful information to distinguish and classify two similar bacterial crotonase superfamily enzymes.     

Sequence analysis of the first B5 subgenogroup strain of enterovirus 71 isolated in Korea

Journal of Microbiology - Enterovirus A71 (EV71), the main etiological agent of handfoot- mouth disease (HFMD), circulates in many areas of the world and has caused large epidemics since 1997,...     

The functional study of human proteins using humanized yeast

Journal of Microbiology - The functional and optimal expression of genes is crucial for survival of all living organisms. Numerous experiments and efforts have been performed to reveal the...     

Practical methodology for gametophyte proliferation and sporophyte production in green penny fern (Lemmaphyllum microphyllum C. Presl) using mechanical fragmentation

Propagation of gametophytes and sporophytes using mechanical fragmentation has been considered a suitable method for mass production of ferns. This study aimed to develop a practical propagation method for Lemmaphyllum microphyllum C. Presl, which is a fern of significant ornamental and medicinal value. Gametophytes were obtained through in vitro spore germination and used for propagation experiments. The gametophyte was mechanically fragmented using a scalpel into small fragments, which were then used to investigate gametophyte proliferation. In addition, the gametophyte was fragmented using a blender and then used to study sporophyte formation. Optimal proliferation conditions of the gametophyte were determined using Murashige and Skoog (MS) basal medium (double-, full-, half-, quarter-strength), Knop medium, and medium components (sucrose, nitrogen sources, activated charcoal), at various concentrations. The fresh weight of the gametophyte was 14-fold higher than that of gametophytes (300 mg) used as culture material, when cultured on double-strength MS. Moreover, 1 g of the gametophyte fragmented in 25 mL of distilled water formed more than 430 sporophytes in a soil mixture in an area of 7.5 cm². The sporophytes were successfully cultivated in the greenhouse after acclimation. A large-scale production method for L. microphyllum that can be easily implemented in a fern production farm is outlined.

     

PICH regulates the abundance and localization of SUMOylated proteins on mitotic chromosomes

Proper chromosome segregation is essential for faithful cell division and if not maintained results in defective cell function caused by the abnormal distribution of genetic information. Polo-like kinase 1–interacting checkpoint helicase (PICH) is a DNA translocase essential for chromosome bridge resolution during mitosis. Its function in resolving chromosome bridges requires both DNA translocase activity and ability to bind chromosomal proteins modified by the small ubiquitin-like modifier (SUMO). However, it is unclear how these activities cooperate to resolve chromosome bridges. Here, we show that PICH specifically disperses SUMO2/3 foci on mitotic chromosomes. This PICH function is apparent toward SUMOylated topoisomerase IIα (TopoIIα) after inhibition of TopoIIα by ICRF-193. Conditional depletion of PICH using the auxin-inducible degron (AID) system resulted in the retention of SUMO2/3-modified chromosomal proteins, including TopoIIα, indicating that PICH functions to reduce the association of these proteins with chromosomes. Replacement of PICH with its translocase-deficient mutants led to increased SUMO2/3 foci on chromosomes, suggesting that the reduction of SUMO2/3 foci requires the remodeling activity of PICH. In vitro assays showed that PICH specifically attenuates SUMOylated TopoIIα activity using its SUMO-binding ability. Taking the results together, we propose a novel function of PICH in remodeling SUMOylated proteins to ensure faithful chromosome segregation.     

Expression Analyses of MicroRNAs in Hamster Lung Tissues Infected by SARS-CoV-2

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is an infectious disease with multiple severe symptoms, such as fever over 37.5°C, cough, dyspnea, and pneumonia. In our research, microRNAs (miRNAs) binding to the genome sequences of severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory-related coronavirus (MERS-CoV), and SARS-CoV-2 were identified by bioinformatic tools. Five miRNAs (hsa-miR-15a-5p, hsa-miR-15b-5p, hsa-miR-195-5p, hsa-miR-16-5p, and hsa-miR-196a-1-3p) were found to commonly bind to SARS-CoV, MERS-CoV, and SARS-CoV-2. We also identified miRNAs that bind to receptor proteins, such as ACE2, ADAM17, and TMPRSS2, which are important for understanding the infection mechanism of SARS-CoV-2. The expression patterns of those miRNAs were examined in hamster lung samples infected by SARS-CoV-2. Five miRNAs (hsa-miR-15b-5p, hsa-miR-195-5p, hsa-miR-221-3p, hsa-miR-140-3p, and hsa-miR-422a) showed differential expression patterns in lung tissues before and after infection. Especially, hsa-miR-15b-5p and hsa-miR-195-5p showed a large difference in expression, indicating that they may potentially be diagnostic biomarkers for SARS-CoV-2 infection.     

CaСl2 Salt Signaling in Primary Root Architecture and Lateral Root Emergence in Arabidopsis thaliana

Russian Journal of Plant Physiology - Plant root architecture modulates during developmental stages and adjusts with the environmental condition. The cytosolic calcium which is a ubiquitous...     

Insight into the bovine milk peptide LPcin‐YK3 selection in the proteolytic system of Lactobacillus species

Antimicrobial peptides are class of small, positively charged peptides known for their broad‐spectrum antimicrobial activity. Antimicrobial activities for most antimicrobial peptides have largely remained elusive, particularly in the lactic acid bacteria. However, recently our investigation using LPcin‐YK3, an antimicrobial peptide from bovine milk, suggests that in vitro antimicrobial activity was reduced over 100‐fold compared with pathogenic bacteria. Additionally, for the structural study of how antimicrobial peptide undergoes its reaction at the proteolytic pathway of lactic acid bacteria based on degradation assay and propidium iodide staining, we performed molecular docking for interaction between oligopeptide‐binding protein A and LPcin‐YK3 peptide. Given that degradation related to the LPcin‐YK3 peptide in lactic acid bacteria proteolytic system, the inhibitory inactivity of LPcin‐YK3 against beneficial lactic acid bacteria strains may be one of the primary pharmacological properties of recombinant peptide discovered in bovine milk. These results provide structural and functional insights into the proteolytic mechanism and possibility as a putative substrate of oligopeptide‐binding protein A in respect of LPcin‐YK3 peptide.