A secreted phospholipase A2 (PLA2) has been discovered. It is produced by Streptomyces violaceoruber A-2688 and is the first PLA2 identified in prokaryote. In this study, the enzyme is demonstrated to be useful in measuring the content of calcium(II) ion in fluid, such as plasma and serum. Two methods were established for the measurement: one measures the increase of fluorescence intensity liberated as a result of the hydrolysis of phosphatidylcholine carrying a fluorescence-tag using the bacterial PLA2; the other, a spectrophotometric method, is based on the increase in 500-nm absorbance due to quinoneimine-dye formed as a result of the hydrolysis of lecithin by PLA2. These enzymatic measurements are novel methods for the determination of calcium(II) ion content.
We monitored the fate of actinophage Yok-15 isolated from Thai soil, and of its host bacteria, Streptomyces sp. strain 15 (just like Yok-15, both were isolated from Thai soil), S. coelicolor, S. griseus, and S. viridochromogenes in two kinds of Thai and Japanese sterile soil using batch microcosms. Phage Yok-15 had a wide host range and infected 39 important strains of Streptomyces species among the 48 strains tested. In two kinds of Thai soil, phage Yok-15 multiplied to a maximum titer, and then the titer decreased rapidly during incubation with strain 15. However, multiplication of phage YoK-15 was nil during incubation with three other streptomycetes in Thai soil, as well as with all the streptomycetes used in Japanese soil. During incubation with these streptomycetes, the phages decreased more rapidly in titer than did free phages in both Thai and Japanese soil. Therefore, the nature or conditions of the soil and host streptomycetes used affected the survival and multiplication of phages in soil. On the other hand, the survival of host streptomycetes seemed to be more strongly affected by conditions of soil rather than by phage infection.
An extracellular polysaccharide (EPS) prepared from SM-1, a mucoidal mutant of Rhodococcus rhodochrous ATCC 12674 was found to absorb and retain moisture in both dry and high-temperature environments. The EPS absorbed more than 120% and 17% of its weight in moisture under the conditions of 32% and 11% relative humidity, respectively, representing much higher absorption than that of known moisture absorbents such as silica gel, glycerol, and hyaluronic acid. The SM-1 EPS is an acidic polysaccharide containing D-galactose, D-glucose, L-fucose, and D-glucuronic acid at a molar ratio of 6: 3: 2: 4. In addition, 1.2% (W/W) stearic acid, 2.3% (W/W) palmitic acid, and 10.3% (W/W) pyruvic acid were also contained in the SM-1 EPS. These evidences indicate that the SM-1 EPS is a novel moisture-absorbing biopolymer.
We found novel relC (=rplK) mutants from the Streptomyces coelicolor A3(2) strain. The mutant strains displayed a point mutation or a deletion mutation within the rplK gene, which codes for the ribosomal L11 protein. These relC mutants exhibited a markedly reduced ability to accumulate ppGpp upon nutritional shift-down and showed an inability to produce the antibiotic actinorhodin, although the mutants grew as well as the parental strain. Gene replacement experiments revealed that the relC mutation alone is responsible for this impaired ability to produce actinorhodin. This failure to produce the actinorhodin phenotype is conditional, as the relC mutants were capable of producing actinorhodin when cultured on plates containing a limited amount of phosphate.
The nucleotide sequence of the dnaK locus in Streptomyces griseus 2247 revealed a gene organization consisting of 6 genes (5’-orf1-dnaK-grpE-dnaJ-hspR-orf6-3’). The genes orf1 and orf6 were predicted to transcribe in the direction opposite that of dnaK, grpE, dnaJ and hspR. This locus of S. griseus was compared with that of S. coelicolor A3(2). No homolog of orf1 and orf6 was found in S. coelicolor, while dnaK, grpE, dnaJandhspR were highly similar to those of S. coelicolor. Promoter probing analysis revealed significant promoter activity in the upstream region of the dnaK gene. The interspace regions of contiguous genes (dnaK-grpE-dnaJ-hspR) were amplified by RT-PCR of mRNA, indicating that these four genes constitute a polycistronic operon. His-tagged HspR protein was overexpressed in Escherichia coli and purified.
Three applications of engineered polyketide synthase genes are reviewed in which the erythromycin and FK506 genes were used to make novel antibacterial and neuroregenerative drugs. Heterologous expression of the genes for industrial production of antibacterial and anticancer drugs is also featured.