Biology subjects

Masuda, T.

Publications and source records attributed to Masuda, T..

3 recordsLinked to original sources

bioRxiv2018

Homologous expression and characterization of gassericin T and gassericin S, a novel class IIb bacteriocin produced by Lactobacillus gasseri LA327

Lactobacillus gasseri LA327 isolated from the large intestine tissue in humans is a bacteriocinogenic strain and is predicted to produce two kinds of class IIb bacteriocins, i.e. gassericin T (GT) and acidocin LF221A (Acd LF221A). In this study, DNA sequencing of the genes for GT and Acd LF221A on Lb. gasseri LA327 revealed that the amino acid sequences for GT completely corresponded with those of gat except for GatK (histidine kinase). However, those for the Acd LF221A had analogues which differed in at least one amino acid residue to be a putative class IIb bacteriocin designated as gassericin S (GS). By deletion test of GT structural genes (gatAX), the LA327 strain retained the bacteriocin activity, and the LA327 mutant strain lacking the ABC-type transporter gene (gatT) completely lost the bacteriocin activity. This indicates that LA327 strain is a GS producer, and GS production is performed via gat with the inclusion of gatT. Homologous expression using deletion mutants for GS and GT containing each single peptide elucidated that GS (GasAX) and GT (GatAX) showed synergistic activity as class IIb bacteriocins, respectively, and no synergistic activity was observed between each peptide of GS and GT. The molecular mass of GS was estimated to be theoretical ca. 5,400 Da by in situ activity assay after SDS-PAGE, clarifying that GS was actually expressed as an active class IIb bacteriocin. Furthermore, stability of GS expressed against pH, heat and protease was determined.\n\nImportanceWe determined the complete DNA sequence for GS, a novel class IIb bacteriocin of Lb. gasseri, and succeeded to express GS as active bacteriocins. Our results clarified the interaction of each class IIb component peptide for GT in addition to GS via construction of homologous mutants which were not dependent on the purification. These data may demonstrate the characteristics of class IIb bacteriocins for Lb. gasseri.

microbiology↗

bioRxiv2018

Cell competition corrects noisy Wnt morphogen gradients to achieve robust patterning

Morphogen signaling forms an activity gradient and instructs cell identities in a signaling strength-dependent manner to pattern developing tissues. However, developing tissues also undergo dynamic morphogenesis, which may produce cells with unfit morphogen signaling and consequent noisy morphogen gradient. Here we show that a cell competition-related system corrects such noisy morphogen gradients. Zebrafish imaging analyses of the Wnt/{beta}-catenin signaling-gradient, which acts as a morphogen to establish embryonic anterior-posterior patterning, revealed that unfit cells with abnormal Wnt/{beta}-catenin activity spontaneously appear and produce noise in the Wnt/{beta}-catenin-gradient. Communication between the unfit and neighboring fit cells via cadherin proteins stimulates the apoptosis of the unfit cells by activating Smad signaling and reactive oxygen species production. This unfit cell elimination is required for proper Wnt/{beta}-catenin-gradient formation and consequent anterior-posterior patterning. Because this gradient controls patterning not only in the embryo but also in adult tissues, this system may support tissue robustness and disease prevention.

cell biology↗

bioRxiv2017

Atomic resolution structure of serine protease proteinase K at ambient temperature

Atomic resolution structures (beyond 1.20 [A]) at ambient temperature, which is usually hampered by the radiation damage in synchrotron X-ray crystallography (SRX), will add to our understanding of the structure-function relationships of enzymes. Serial femtosecond crystallography (SFX) has attracted surging interest by providing a route to bypass such challenges. Yet the progress on atomic resolution analysis with SFX has been rather slow. In this report, we describe the 1.20 [A] resolution structure of proteinase K using 13 keV photon energy. Hydrogen atoms, water molecules, and a number of alternative side-chain conformations have been resolved. The increase in the value of B-factor in SFX suggests that the residues and water molecules adjacent to active sites were flexible and exhibited dynamic motions at specific substrate-recognition sites.

biochemistry↗