Two groups control light-induced schiff base deprotonation and the proton affinity of asp(85) in the Arg(82)His mutant of bacteriorhodopsin

Arg(82) is one of the four buried charged residues in the retinal binding pocket of bacteriorhodopsin (bR). Previous studies show that Arg(82) controls the pK(a)s of Asp(85) and the proton release group and is essential for fast light-induced proton release. To further investigate the role of Arg(82) in light-induced proton pumping, we replaced Arg(82) with histidine and studied the resulting pigment and its photochemical properties. The main pK(a) of the purple-to-blue transition (pK(a) of Asp(85)) is unusually low in R82H: 1.0 versus 2.6 in wild type (WT). At pH 3, the pigment is purple and shows light and dark adaptation, but almost no light-induced Schiff base deprotonation (formation of the M intermediate) is observed. As the pH is increased from 3 to 7 the M yield increases with pK(a) 4.5 to a value approximately 40% of that in the WT. A transition with a similar pK(a) is observed in the pH dependence of the rate constant of dark adaptation, k(da). These data can be explained, assuming that some group deprotonates with pK(a) 4.5, causing an increase in the pK(a) of Asp(85) and thus affecting k(da) and the yield of M. As the pH is increased from 7 to 10.5 there is a further 2.5-fold increase in the yield of M and a decrease in its rise time from 200 &mgr;s to 75 &mgr;s with pK(a) 9. 4. The chromophore absorption band undergoes a 4-nm red shift with a similar pK(a). We assume that at high pH, the proton release group deprotonates in the unphotolyzed pigment, causing a transformation of the pigment into a red-shifted "alkaline" form which has a faster rate of light-induced Schiff base deprotonation. The pH dependence of proton release shows that coupling between Asp(85) and the proton release group is weakened in R82H. The pK(a) of the proton release group in M is 7.2 (versus 5.8 in the WT). At pH < 7, most of the proton release occurs during O --> bR transition with tau approximately 45 ms. This transition is slowed in R82H, indicating that Arg(82) is important for the proton transfer from Asp(85) to the proton release group. A model describing the interaction of Asp(85) with two ionizable residues is proposed to describe the pH dependence of light-induced Schiff base deprotonation and proton release.     

A new species of Scorzonera L. (Asteraceae) from south Anatolia, Turkey

A new species Scorzonera gokcheoglui O. Ünal & R. S. Göktürk sp. nov. from south Anatolia is described and illustrated. Its relationships with S. argyria and S. pisidica are discussed. A map showing the distribution of the species and other related species is given.  © 2003 The Linnean Society of London, Botanical Journal of the Linnean Society , 2003, 142 , 465–468.     

Complete intra-annual cycle of leaf area index in a Platanus orientalis L. stand

Leaf area index (LAI) analysis of deciduous forest trees is usually restricted to seasonal monitoring involving the assessment of distinct leaf phenological stages within definite time intervals of the year. However, continuous LAI monitoring that includes entire leaf periods is necessary to define the ecophysiological characteristics of deciduous trees. Therefore, this study investigated the intra-annual cycle of the LAI for a Platanus orientalis L. stand in the Bart?n watershed of Turkey. A complete cycle involves three periods: foliation, stable, and defoliation. The foliation period comprises budburst, leaf emergence and flushing sessions, whereas the defoliation period consists of leaf senescence and leaf fall sessions. The stable period is in between these two periods when LAI values are at a climax around maximum. Eight points were determined in the field for the analysis of LAI by a hemispherical photography technique. Over a relatively frequent schedule, photographs were taken almost weekly during the foliation period. Both weekly and approximate monthly photographs were applied during the stable period. Finally, near-monthly photographs were taken for the defoliation period. The foliation period lasted for about 1.5 months from mid-April to May with the mean LAI reaching from 0.16 up to 2.38. Mean LAI was between 2.38 and 2.47 for a stable period over 2 months (June and July). For the defoliation period, mean LAI dropped from 2.42 down to 0.35 over 5 months from August to December. The total foliated period was more than 8 months, which is relatively long for a temperate forest. In addition, correlations between mean LAI and maximum, mean and minimum temperatures were highly significant (P < 0.01) with coefficients (r) of 0.79, 0.90 and 0.93, respectively. By describing the intra-annual LAI pattern, this study fills a gap in the literature on the phenology of Platanus orientalis L.     

The environmental significance of algae in the mi Llandovery succession of the central Oslo Region

Girvanella , micritization phenomena and the dasycladacean alga Cyclocrinites are found in the middle Llandovery succession of the central Oslo Region. Both Girvanella and micritization phenomena occur in carbonate-dominated sequences where sedimentary structures and associated faunas suggest depositional environments above normal wave base. In contrast, Cyclocrinites occurs in shale-dominated sequences where other evidence suggests quiet, deep-water environments. We suggest that Cyclocrinites would not normally be preserved in situ in its life environment in relatively turbulent shallow waters. Preservation most commonly resulted from post mortem transport into low-energy muddy environments devoid of in situ algal activity. The resultant fossil is a flattened cast of this originally spherical alga.     

Quest for wine yeasts—An old story revisited

Numerous studies have described the yeast biota of grapes, and grape must in order to understand better the succession of yeasts during fermentation of wine. The origin of the wine yeasts has been rather controversial. By using more elaborate isolation methods, classical genetic analysis and electrophoretic karyotyping of monosporic clones, with this study, credible proof now exists that the vineyard is the primary source for the wine yeasts and that strains found on the grapes can be followed through the fermentation process.     

Molecular population genetics of the alcohol dehydrogenase locus in the Hawaiian drosophilid D. mimica

Sequence variation among 10 alleles of the alcohol dehydrogenase (Adh) gene of the Hawaiian drosophilid D. mimica was analyzed with reference to the evolutionary history of the Hawaiian subgroup as well as to levels and patterns of polymorphism of the Adh gene in continental drosophilid species. The Adh gene of D. mimica is less polymorphic than that of other drosophilid species, and no replacement substitutions were found. Statistical analyses of the Adh alleles suggested the action of balancing selection and revealed significant linkage disequilibrium among three of the variable sites. The effective population size was estimated to be only slightly smaller than that of continental species and, surprisingly, on the same order of magnitude as the actual size.      

Cloning, expression, purification, and characterization of the acid alpha-mannosidase from Trypanosoma cruzi

The acid alpha-mannosidase of Trypanosoma cruzi is a broad-specificity hydrolase involved in the catabolism of glycoconjugates, presumably in the digestive vacuole. We have cloned the alpha-mannosidase gene from a T.cruzi epimastigote genomic library. The alpha-mannosidase gene was determined to be single copy by Southern analysis, and similar sequences were not detected in genomic digests of either Trypanosoma brucei or Leishmania donovani. The coding region was subcloned into the Pichia pastoris expression vector pPICZ, and alpha-mannosidase activity was detected in the medium of induced cultures. The recombinant alpha- mannosidase demonstrated a pH optimum, inhibition by swainsonine, Km, and substrate specificity consistent with the characteristics of the alpha-mannosidase previously purified from T.cruzi epimastigotes. The recombinant enzyme was purified 103-fold from the culture medium of Pichia pastoris and had a native molecular mass of 359 kDa by gel filtration. A combination of SDS-PAGE, deglycosylation with endo H, and NH2-terminal sequencing indicates that the enzyme is originally synthesized as a homodimeric polypeptide that is subsequently cleaved to form a heterotetramer composed of 57 and 46 kDa subunits. A polyclonal antibody raised to the recombinant enzyme was shown to immunoprecipitate the alpha-mannosidase from T.cruzi cell extracts and will be used in future immunolocalization studies.