Application of halophilic nuclease H of Micrococcus varians subsp. halophilus to commercial production of flavoring agent 5'-GMP

RNA was degraded at 60 degrees C for 24 h by halophilic nuclease H in supernatants from broth cultures of Micrococcus varians subsp. halophilus containing 12% NaCl. Since contaminating 5'-nucleotidase exhibited almost no activity under these conditions, the 5'-GMP formed could be recovered from the reaction mixture, and the yield was 805 mg from 5 g of RNA.     

A Markov approach for describing post-fire succession of vegetation

Vegetation dynamics in the coastal area of the Seto Inland Sea region in Japan, where wild fires occur frequently, were described using a stationary Markov model. In this region, vegetation types ofMiscanthus-Pleioblastus grassland,Lespedeza-Mallotus scrub,Pinus-Rhododendron forest andCrassocephalum-Erechtites community have been identified, and these show cyclic succession under the influence of fires. The model uses parameters determining fire frequency and rate of successional change to analyze the effect of variation in these parameters on the areal ratio of each vegetation type at equilibrium and on the time taken for one vegetation type to succeed another (elapsed successional time). The effect of fire frequency differs between hypothetical habitats with high and low productivity. A policy for vegetation management in areas of high and low productivity is proposed. The advantages and limitations of applying Markov models to studies of vegetation succession are also discussed.     

Electron microscopic studies on myosin molecules from chicken gizzard muscle III. Myosin dimers

Previously, we (Onishi, H. & Wakabayashi, T. (1982) J. Biochem. 92, 871) reported that the ATP-induced disassembly of chicken gizzard "thick filaments" resulted in myosin monomers with "looped" tails. In the present study, we found that these monomers assembled themselves into antiparallel dimers when they were placed in a medium of low ionic strength (approximately 2 mM).     

Wavelength-dependence of the relative rate constants for the main geometric and structural photoisomerization of bilirubin IX alpha bound to human serum albumin. Demonstration of green light at 510 nm as the most effective wavelength in photochemical changes from (ZZ)-bilirubin IX alpha to (EZ)-cyclobilirubin IX alpha via (EZ)-bilirubin.

The kinetics for the quantitatively important reaction: (Formula: see text) that is, the photochemical interconversion between bilirubin and its geometric and structural photoisomers bound to human serum albumin in aqueous solution when various wavelengths of monochromatic light were used, were assayed by h.p.l.c. In order to clarify the wavelength-dependence of the relative rate constants in the individual steps, a light-source with a half-bandwidth of 10 nm was used at increments of 20 nm, in the range from 410 nm to 550 nm. We describe for the first time studies on the wavelength-dependence of rate constants in geometric and structural photoisomerization reactions in vitro of (ZZ)-bilirubin or (EZ)-bilirubin bound to human serum albumin, especially the relative rate constants of cyclization of (EZ)-bilirubin into (EZ)-cyclobilirubin. Because studies in vitro have demonstrated that the wavelengths from 350 to 450 nm are mutagenic, the results obtained indicated that the safest and ideal light-source for phototherapy is green light of 510 nm, which keeps (ZE)-bilirubin concentrations as low as possible, as shown by a maximal value of k2 at 510 nm and a relatively low value of k1 at 510 nm. This light-source still ensures the substantial absorption of (ZZ)-bilirubin, which is the precursor of (EZ)-bilirubin, the intermediate in (EZ)-cyclobilirubin formation and, furthermore, as shown by the maximal value of k5 and a considerable value of k4 at 510 nm, promotes the cyclization of (EZ)-bilirubin derived from (ZZ)-bilirubin even though k3 at 510 nm also shows a peak value.     

Odorant transfer characteristics of white bread during baking

The potent odorants in the crust and crumb of white bread were identified and quantified by gas chromatography-mass spectrometry and gas chromatography/olfactometry. The weight loss ratio of the samples baked at 220 °C was controlled in the range of 0-28%. The odorants were classified into 5 types by the transfer characteristics: i) All amounts of odorant transferred from the crust to external space (type-I). ii) All transferred from the crust to the crumb and external space (type-II). iii) Certain amount remaining in the crust and the rest transferred to the crumb and external space (type-III). iv) All transferred from the crumb to external space (type-IV). v) Certain amount remaining in the crumb and the rest transferred to the crust and external space (type-V). The odorants of type-IV were not apparent after the crust had formed. The results indicate that the crust could be a barrier to prevent the odorants from being transferred to external space.     

Geranylgeranyltransferase I of Candida albicans: null mutants or enzyme inhibitors produce unexpected phenotypes

Geranylgeranyltransferase I (GGTase I) catalyzes the transfer of a prenyl group from geranylgeranyl diphosphate to the carboxy-terminal cysteine of proteins with a motif referred to as a CaaX box (C, cysteine; a, usually aliphatic amino acid; X, usually L). The alpha and beta subunits of GGTase I from Saccharomyces cerevisiae are encoded by RAM2 and CDC43, respectively, and each is essential for viability. We are evaluating GGTase I as a potential target for antimycotic therapy of the related yeast, Candida albicans, which is the major human pathogen for disseminated fungal infections. Recently we cloned CaCDC43, the C. albicans homolog of S. cerevisiae CDC43. To study its role in C. albicans, both alleles were sequentially disrupted in strain CAI4. Null Cacdc43 mutants were viable despite the lack of detectable GGTase I activity but were morphologically abnormal. The subcellular distribution of two GGTase I substrates, Rho1p and Cdc42p, was shifted from the membranous fraction to the cytosolic fraction in the cdc43 mutants, and levels of these two proteins were elevated compared to those in the parent strain. Two compounds that are potent GGTase I inhibitors in vitro but that have poor antifungal activity, J-109,390 and L-269,289, caused similar changes in the distribution and quantity of the substrate. The lethality of an S. cerevisiae cdc43 mutant can be suppressed by simultaneous overexpression of RHO1 and CDC42 on high-copy-number plasmids (Y. Ohya et al., Mol. Biol. Cell 4:1017, 1991; C. A. Trueblood, Y. Ohya, and J. Rine, Mol. Cell. Biol. 13:4260, 1993). Prenylation presumably occurs by farnesyltransferase (FTase). We hypothesize that Cdc42p and Rho1p of C. albicans can be prenylated by FTase when GGTase I is absent or limiting and that elevation of these two substrates enables them to compete with FTase substrates for prenylation and thus allows sustained growth.     

Functional roles of ionic and hydrophobic surface loops in smooth muscle myosin: their interactions with actin

This investigation ascertains whether, in (smooth muscle) myosin, certain residues engage in functional interactions with their actin conjugates in an actomyosin complex. Such interactions have been postulated from putting together crystallographic models of the two proteins [Rayment, I., Rypniewski, W. R., Schmidt-B?se, K., Smith, R., Tomchick, D. R., Benning, M. M., Winkelmann, D. A., Wesenberg, G., and Holden, H. M. (1993) Science 261, 50-58]. Here, in several instances, we ask whether mutation of a particular residue significantly impairs a function, and find that the answers are largely rationalized by the original postulation. Additionally, a novel element emerges from our investigation. To assess function, we test the wild type and mutant systems as they perform in the steady state of ATP degradation. In doing so, we assume, as usual, that degradation proceeds from an early stage in which the complex forms (and is described by parameter K(app)) to a later stage during which the product leaves the complex (and is described by parameter V(max)). Interestingly, certain defects induced by the mutations are associated with changes in K(app), and other defects are associated with changes in V(max), suggesting that our procedure at least roughly distinguishes between events according to the time in the degradation at which they occur. In this framework, we suggest that (1) in the actin-myosin association phase, cationic residues Lys-576 and Lys-578 interact with anionic residues of the so-called second actin, and (2) in the product leaving phase, hydrophobic residues Trp-546, Phe-547, and Pro-548, as well as the Thr-532/Asn-533/Pro-534/Pro-535 sequence, sever connections with the so-called first actin. The role of Glu-473 is also examined.