A novel marine bacterium, strain KMM 6050T, was isolated from the sea urchin Strongylocentrotus intermedius, which inhabits the Sea of Japan. The strain studied was strictly aerobic, heterotrophic, yellow–orange-pigmented, motile by gliding, Gram-negative and oxidase-, catalase-, β-galactosidase- and alkaline phosphatase-positive. The results of 16S rRNA gene sequence analysis showed that strain KMM 6050T occupies a distinct lineage within the family Flavobacteriaceae and is most closely related to the species Mesonia algae and Salegentibacter salegens (sequence similarity of 92·5–92·6 %). The DNA G+C content of KMM 6050T was 39·6 mol%. The major respiratory quinone was MK-6. The predominant fatty acids were i15 : 0, a15 : 0, 15 : 0, i16 : 1, i16 : 0, i16 : 0 3-OH and i17 : 0 3-OH. On the basis of phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics, the novel bacterium has been assigned to the genus Gramella gen. nov., as Gramella echinicola sp. nov. The type strain is KMM 6050T (=KCTC 12278T=NBRC 100593T=LMG 22585T).

Streptomyces exfoliatus SMF13 produced leupeptin, chymotrypsin-l i ke protease (CTP), metalloprotease, and trypsin-like protease (TLP) extracellularly. The activity of TLP was specifically inhibited by leupeptin. Production of leupeptin was closely associated with growth but leupeptin was inactivated by leupeptin-inactivating protein (LIP) when growth reached the stationary phase in submerged cultures, or when aerial mycelia started to form on surface cultures. Autolysis of mycelia after the stationary phase in submerged cultures was apparently retarded by the addition of leupeptin; on surface cultures, aerial mycelium formation was clearly retarded by the addition of leupeptin. We propose that CTP participates primarily in utilization of a proteinaceous nitrogen source, that TLP functions as an essential enzyme involved in the metabolism of mycelial protein, that leupeptin inhibits the activity of TLP and that LIP inactivates leupeptin. The cascade of regulatory actions of the compounds, which are produced sequentially during mycelium development, may provide selective advantages in adverse culture conditions.

The specific rates of growth, substrate utilization, and ethanol production as well as yields of biomass and ethanol production on xylose for the recombinant Zymomonas mobilis ZM4(pZB5) were shown to be much less than those on glucose or glucose-xylose mixtures. Typical fermentations with ZM4(pZB5) growing on glucosexylose mixtures followed two-phase growth kinetics with the initial uptakes of glucose and xylose being followed by slower growth and metabolic uncoupling on xylose after glucose depletion. The reductions in rates and yields from xylose metabolism were considered in the present investigation and may be due to a number of factors, including the following: (i) the increased metabolic burden from maintenance of plasmid-related functions, (ii) the production of by-products identified as xylitol, acetate, lactate, acetoin, and dihydroxyacetone by 13 Cnuclear magnetic resonance (NMR) spectroscopy and high-performance liquid chromatography, (iii) growth inhibition due to xylitol by the putative inhibitory compound xylitol phosphate, and (iv) the less energized state of ZM4(pZB5). In vivo 31 P-NMR studies have established that the levels of NTP and UDP sugars on xylose were less than those on glucose, and this energy limitation is likely to restrict the growth of the recombinant strain on xylose media.Zymomonas mobilis has attracted widespread interest for fuel ethanol production because of its higher specific rates of sugar uptake and ethanol production, higher ethanol tolerance, and higher ethanol conversion efficiencies when compared to the traditionally used yeasts (10,14,21,22,26). However, wild-type strains of Z. mobilis can only utilize glucose, fructose, and sucrose, and they lack the pentose metabolism pathway necessary to ferment such sugars as xylose or arabinose. The cloning of enzymes for xylose assimilation and metabolism in Z. mobilis has now been reported (30), and a subsequent study has resulted in the successful integration of the requisite genes into the Zymomonas genome (M. Zhang, Y. C. Chou, X. K. Lai, S. Milstrey, N. Danielson, K. Evans, A. Mohagheghi, and M. Finkelstein, Abstr. 21st Symp. Biotechnol. Fuels Chem., abstr. 2-16, 1999). These genetically engineered strains can now convert xylose to ethanol by the combined use of the Entner-Doudoroff and pentose pathways facilitated by the cloned enzymes xylose isomerase and xylulokinase for xylose assimilation and by transketolase and transaldolase for pentose metabolism. In a further study, the cloning of three additional enzymes for arabinose utilization has been reported (4). However, when xylose is the sole carbon source, lower biomass yields and slower growth rates as well as lower ethanol yields for recombinant strains have been reported (9, 11, 12, 23; H. G. Lawford and J. D. Rousseau, Abstr. 21st Symp. Biotechnol. Fuels Chem., abstr. 2-24, 1999).In this study, the fermentation characteristics of the recombinant Z. mobilis ZM4(pZB5) on xylose or glucose alone, or on xylose-glucose mixtures, have been investigated to determine possib...

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