Role of Acetyl CoA: Deacetylcephalosporin C Acetyltransferase in Cephalosporin C Biosynthesis by Cephalosporium acremonium

Existence of an acetyltransferase, which catalizes acetylation of deacetylcephalosporin C to cephalosporin C, was demonstrated for the first time in cell-free extracts of Cephalosporium acremonium. The pH optimum of the enzyme appeared to be 7.0 to 7.5 and the enzyme required essentially Mg²⁺ as a cofactor for its reaction. The activity of this enzyme was not observed in the cell-free extracts of deacetylcephalosporin C-producing mutants Nos. 20, 29, 36 and 40, but was recovered in a revertant obtained from the mutant No. 40. These results indicate that deacetylcephalosporin C accumulation by these mutants was due to the lack of the acetyltransferase and made it reasonable that the terminal reaction of cephalosporin C biosynthesis in Cephalosporium acremonium proceeded by the catalytic action of acetyltransferase.     

Inhibition of malaria parasite growth by quinomycin A and its derivatives through DNA-intercalating activity

Quinomycin A and its derivatives were identified as potent antimalarial (Plasmodium falciparum) agents in a screen of the RIKEN NPDepo chemical library. IC₅₀ values of quinomycin A and UK-63,598 were approximately 100 times lower than that of the antimalarial drug chloroquine. This activity was mitigated by the addition of plasmid DNA, suggesting that these compounds act against parasites by intercalating into their DNA.     

Purification of Cytochrome a of an L-Glutamate-producing Microorganism,Brevibacterium thiogenitalis

The respiratory chain system of Brev. thiogenitalis grown in the presence of copper ions contained cytochromes a, b and c. The cytochrome a was solubilized and purified from the cell-free extracts by means of Triton X-100 and cholate extraction, and DEAE-cellulose chromatography. It was purified about 130-fold from the cell-free extracts and was free from other cytochromes, The purified preparation contained 1.4 mμatom copper and 1.9 mμatom iron per mμmole heme a, respectively, and approximately 5 mμmoles heme a per mg protein.     

Role of Copper Ions in the Regulation of L-Glutamate Biosynthesis

Cell-free extracts of Brevibacterium thiogenitalis culture grown in the presence of copper catalyzed the oxidation of NADH₂ and succinate through an electron transport chain which contained menaquinones and cytochromes a, b and c. On the other hand, extracts of cells grown in the absence of copper lacked cytochromes a and c, and contained cytochrome d.

These findings, as well as the results obtained in inhibition experiments, suggest that in copper-deficient cells the major part of NADH₂ was oxidized via a bypass in which the electrons were transferred directly from flavoprotein or cytochrome b to molecular oxygen.

Electron transport from these substrates to molecular oxygen resulted in ATP synthesis. The average P/O ratios in extracts of the copper-sufficient cells were 0.33 for generated NADH₂, 0.20 for added NADH₂, and 0.34 for succinate, and those in extracts of the copper-deficient cells were 0.15, 0.13 and 0.21, respectively. In addition, a linear relationship was found between the yield of L-glutamate from acetate and the P/Ο ratios with both NADH₂ and succinate as substrates.

From these results, it is reasonable to consider that the poor yield of L-glutamate from acetate in copper-deficient cells was due to a reduction in energy supply, which was caused by the low efficiency of oxidative phosphorylation.     

Bifunctional properties and characterization of a novel sialidase with esterase activity from Bifidobacterium bifidum

ABSTRACT

Sialidases catalyze the removal of terminal sialic acid from various complex carbohydrates. In the gastrointestinal tract, sialic acid is commonly found in the sugar chain of mucin, and many enteric commensals use mucin as a nutrient source. We previously identified two different sialidase genes in Bifidobacterium bifidum, and one was cloned and expressed as an extracellular protein designated as exo-α-sialidase SiaBb2. The other exo-α-sialidase gene (siabb1) from the same bifidobacterium encodes an extracellular protein (SiaBb1) consisting of 1795 amino acids with a molecular mass of 189 kDa. SiaBb1 possesses a catalytic domain that classifies this enzyme as a glycoside hydrolase family 33 member. SiaBb1 preferentially hydrolyzes α2,3-linked sialic acid over α2,6-linked sialic acid from sialoglycan, which is the same as SiaBb2. However, SiaBb1 has an SGNH hydrolase domain with sialate-O-acetylesterase activity and an N-terminal signal sequence and C-terminal transmembrane region. SiaBb1 is the first bifunctional sialidase identified with esterase activity.

Abbreviations: GalNAc: N-acetyl-D-galactosamine; Fuc: L-fucose; Gal: D-galactose     

Replication Study in a Japanese Population of Six Susceptibility Loci for Type 2 Diabetes Originally Identified by a Transethnic Meta-Analysis of Genome-Wide Association Studies

AimWe performed a replication study in a Japanese population to evaluate the association between type 2 diabetes and six susceptibility loci (TMEM154, SSR1, FAF1, POU5F1, ARL15, and MPHOSPH9) originally identified by a transethnic meta-analysis of genome-wide association studies (GWAS) in 2014.MethodsWe genotyped 7,620 Japanese participants (5,817 type 2 diabetes patients and 1,803 controls) for each of the single nucleotide polymorphisms (SNPs) using a multiplex polymerase chain reaction invader assay. The association of each SNP locus with the disease was evaluated using logistic regression analysis.ResultsOf the six SNPs examined in this study, four (rs6813195 near TMEM154, rs17106184 in FAF1, rs3130501 in POU5F1 and rs4275659 near MPHOSPH9) had the same direction of effect as in the original reports, but two (rs9505118 in SSR1 and rs702634 in ARL15) had the opposite direction of effect. Among these loci, rs3130501 and rs4275659 were nominally associated with type 2 diabetes (rs3130501; p = 0.017, odds ratio [OR] = 1.113, 95% confidence interval [CI] 1.019–1.215, rs4275659; p = 0.012, OR = 1.127, 95% CI 1.026–1.238, adjusted for sex, age and body mass index), but we did not observe a significant association with type 2 diabetes for any of the six evaluated SNP loci in our Japanese population.ConclusionsOur results indicate that effects of the six SNP loci identified in the transethnic GWAS meta-analysis are not major among the Japanese, although SNPs in POU5F1 and MPHOSPH9 loci may have some effect on susceptibility to type 2 diabetes in this population.     

Dnajb8, a Member of the Heat Shock Protein 40 Family Has a Role in the Tumor Initiation and Resistance to Docetaxel but Is Dispensable for Stress Response

Cancer stem-like cells (CSCs)/cancer-initiating cells (CICs) are defined by their abilities of tumor initiation, self-renewal and differentiation. In a previous study, we showed by gene knockdown using siRNA and gene overexpression experiments that Dnaj (Hsp40) homolog, subfamily B, member 8 (DNAJB8), a role in the maintenance, of renal cell carcinoma CSCs/CICs. In the present study, we established Dnajb8 knockout (KO) renal cell carcinoma (RCC) line cells (RenCa cells) and analyzed the cells to confirm the function of Dnajb8 in RCC CSCs/CICs. Dnajb8 KO cells showed reduced ratios of side population cells and reduced sphere forming ability. An in vivo single cell tumor initiation assay revealed that the numbers of CSCs/CICs were 3 in 4 wild-type RenCa cells and 1 in 4 Dnajb8 KO cells. Dnajb8 KO cells showed sensitivity to Docetaxel. On the other hand, Dnajb8 KO cells did not show any sensitivities to stresses including low pH, low glucose, heat shock and sensitivity to cisplatin. The results indicate that Dnajb8 has a role in tumor initiation, side population ratio and sphere formation but it is dispensable for stress responses.