α-1,3-Glucanase from Streptomyces thermodiastaticus HF3-3 (Agl-ST) has been classified in the glycoside hydrolase (GH) family 87. Agl-ST is a multi-modular domain consisting of an N-terminal β-sandwich domain (β-SW), a catalytic domain, an uncharacterized domain (UC), and a C-terminal discoidin domain (DS). Although Agl-ST did not hydrolyze α-1,4-glycosidic bonds, its amino acid sequence is more similar to GH87 mycodextranase than to α-1,3-glucanase. It might be categorized into a new subfamily of GH87. In this study, we investigated the function of the domains. Several fusion proteins of domains with green fluorescence protein (GFP) were constructed to clarify the function of each domain. The results showed that β-SW and DS domains played a role in binding α-1,3-glucan and enhancing the hydrolysis of α-1,3-glucan. The binding domains, β-SW and DS, also showed binding activity toward xylan, although it was lower than that for α-1,3-glucan. The combination of β-SW and DS domains demonstrated high binding and hydrolysis activities of Agl-ST toward α-1,3-glucan, whereas the catalytic domain showed only a catalytic function. The binding domains also achieved effective binding and hydrolysis of α-1,3-glucan in the cell wall complex of Schizophyllum commune.
Musty odor production by actinomycetes is usually related to the presence of geosmin and 2-methylisoborneol (2-MIB), which are synthesized by enzymes encoded by the geoA and tpc genes, respectively. Streptomyces spp. strain S10, which was isolated from a water reservoir in Malaysia, has the ability to produce geosmin when cultivated in a basal salt (BS) solid medium, but no 2-MIB production occurred during growth in BS medium. Strain S10 could produce higher levels of geosmin when the phosphate concentration was limited to 0.05 mg/L, with a yield of 17.53 ± 3.12 ✕ 105 ng/L, compared with growth in BS medium. Interestingly, 2-MIB production was suddenly detected when the nitrate concentration was limited to 1.0 mg/L, with a yield of 1.4 ± 0.11 ✕ 105 ng/L. Therefore, it was concluded that phosphate- and nitrate-limiting conditions could induce the initial production of geosmin and 2-MIB by strain S10. Furthermore, a positive amplicon of geoA was detected in strain S10, but no tpc amplicon was detected by PCR analysis. Draft genome sequence analysis showed that one open reading frame (ORF) contained a conserved motif of geosmin synthase with 95% identity with geoA in Streptomyces coelicolor A3 (2). In the case of the tpc genes, it was found that one ORF showed 23% identity to the known tpc gene in S. coelicolor A3(2), but strain S10 lacked one motif in the N-terminus.
The cellular fatty acid composition of Aureispira marina IAM 15389T (JCM 23197T), a gliding bacterium isolated from the coastline of Thailand, was re-examined by using a standard MIDI method based on alkaline hydrolysis, and two other methods. The direct transesterification using 5% HCl/methanol or 4 M HCl hydrolysis followed by methyl esterification revealed that 2-hydroxy-15-methyl-hexadecanoic acid (2-OH-iso-C17:0) and 2-hydroxy-15-methyl-hexadecenoic acid (2-OH-iso-C17:1), which were not reported in a previous paper, were found to be major cellular fatty acids of this bacterium, and the amount of 2-OH-iso-C17:1 was even higher than that of arachidonic acid (C20:4), a characteristic polyunsaturated fatty acid present in this bacterium. These 2-hydroxy-fatty acids were contained in two cellular lipids that were relatively stable against alkaline hydrolysis. One of them was analyzed by mass spectrometry, 1H-nuclear magnetic resonance, and other chemical methods, and identified as a ceramide composed of 2-hydroxy-fatty acid and sphingosine of 19 carbons with three double bonds. A minor ceramide containing 18 carbon sphingosine with three double bonds was also detected.
A pot experiment was conducted with kiwifruit planting soil to evaluate the impacts of potassium solubilizing bacteria (KSB) and K-feldspar on the soil nutrient levels, enzyme activities, and microecological environment. The effects were investigated of three inoculation treatments (T1: K-feldspar, T2: KSB, and T3: KSB with K-feldspar) and a non-inoculation treatment (CK) on the enzyme activities and the metabolic activities of the bacterial communities in kiwifruit rhizosphere soil. The results showed that the total nitrogen, available phosphorus, available potassium, and organic matter contents in T3 were 18.19%, 45.22%, 15.06%, and 4.17% higher, respectively, than those in CK at the end of the experiment (90 days). Compared with CK, T3 significantly increased the invertase, urease, acid phosphatase, and polyphenol oxidase activities. T3 had a higher kiwifruit root activity, but there were no significant differences among the four treatments (P > 0.05). T3 significantly altered the bacterial community diversity, increased the utilization of phenolic compounds and polymers, and decreased the utilization of amino acids. Redundancy analysis indicated that soil nutrients (total nitrogen, available phosphorus, and available potassium) and enzyme activities (urease and acid phosphatase) had more important effects on the metabolic activities of the bacterial communities. Co-inoculation enhanced the soil nutrients, enzyme activities, and bacterial community diversity. KSB co-inoculated with K-feldspar has the potential to improve the soil fertility, microbial metabolic activity and plant growth.
Two Indonesian fungi Aspergillus assiutensis BioMCC-f.T.7495 and Penicillium pedernalense BioMCC-f.T.5350 along with a Japanese fungus Hypomyces pseudocorticiicola FKI-9008 have been found to produce gentisyl alcohol (1), which inhibits Plasmodium falciparum dihydroorotate dehydrogenase (PfDHODH) with an IC50 value of 3.4 μM. Another Indonesian fungus, Penicillium citrinum BioMCC-f.T.6730, produced an analog of 1, homogentisic acid (4), which also inhibits PfDHODH with an IC50 value of 47.6 μM.
We selected 96 genes of Aspergillus luchuensis for the construction of a transcription factor gene deletion library. Of these, we successfully deleted 93 genes using Agrobacterium-mediated transformation (AMT) of A. luchuensis RIB 2604 ΔligD strains. We obtained only heterokaryonic strains after deletions of adaB, anBH1, hacA, hapB, hsf1, metR, and sonC gene, and additionally, could not obtain deletion strains for genes abaA and mcmA. The deletion strains will be available through our website (https://www.nrib.go.jp).