New lessons for combinatorial biosynthesis from myxobacteria. The myxothiazol biosynthetic gene cluster of Stigmatella aurantiaca DW4/3-1

The biosynthetic mta gene cluster responsible for myxothiazol formation from the fruiting body forming myxobacterium Stigmatella aurantiaca DW4/3-1 was sequenced and analyzed. Myxothiazol, an inhibitor of the electron transport via the bc(1)-complex of the respiratory chain, is biosynthesized by a unique combination of several polyketide synthases (PKS) and nonribosomal peptide synthetases (NRPS), which are activated by the 4'-phosphopantetheinyl transferase MtaA. Genomic replacement of a fragment of mtaB and insertion of a kanamycin resistance gene into mtaA both impaired myxothiazol synthesis. Genes mtaC and mtaD encode the enzymes for bis-thiazol(ine) formation and chain extension on one pure NRPS (MtaC) and on a unique combination of PKS and NRPS (MtaD). The genes mtaE and mtaF encode PKSs including peptide fragments with homology to methyltransferases. These methyltransferase modules are assumed to be necessary for the formation of the proposed methoxy- and beta-methoxy-acrylate intermediates of myxothiazol biosynthesis. The last gene of the cluster, mtaG, again resembles a NRPS and provides insight into the mechanism of the formation of the terminal amide of myxothiazol. The carbon backbone of an amino acid added to the myxothiazol-acid is assumed to be removed via an unprecedented module with homology to monooxygenases within MtaG.     

Inhibition of biofouling by marine microorganisms and their metabolites

Development of microbial biofilms and the recruitment of propagules on the surfaces of man-made structures in the marine environment cause serious problems for the navies and for marine industries around the world. Current antifouling technology is based on the application of toxic substances that can be harmful to the natural environment. For this reason and the global ban of tributyl tin (TBT), there is a need for the development of "environmentally-friendly" antifoulants. Marine microbes are promising potential sources of non-toxic or less-toxic antifouling compounds as they can produce substances that inhibit not only the attachment and/or growth of microorganisms but also the settlement of invertebrate larvae and macroalgal spores. However, so far only few antilarval settlement compounds have been isolated and identified from bacteria. In this review knowledge about antifouling compounds produced by marine bacteria and diatoms are summarised and evaluated and future research directions are highlighted.     

Development and laboratory-scale testing of a fully automated online flow cytometer for drinking water analysis

Accurate and sensitive online detection tools would benefit both fundamental research and practical applications in aquatic microbiology. Here, we describe the development and testing of an online flow cytometer (FCM), with a specific use foreseen in the field of drinking water microbiology. The system incorporated fully automated sampling and fluorescent labeling of bacterial nucleic acids with analysis at 5-min intervals for periods in excess of 24 h. The laboratory scale testing showed sensitive detection (< 5% error) of bacteria over a broad concentration range (1 × 10(3) -1 × 10(6) cells mL(-1) ) and particularly the ability to track both gradual changes and dramatic events in water samples. The system was tested with bacterial pure cultures as well as indigenous microbial communities from natural water samples. Moreover, we demonstrated the possibility of using either a single fluorescent dye (e.g., SYBR Green I) or a combination of two dyes (SYBR Green I and Propidium Iodide), thus broadening the application possibilities of the system. The online FCM approach described herein has considerable potential for routine and continuous monitoring of drinking water, optimization of specific drinking water processes such as biofiltration or disinfection, as well as aquatic microbiology research in general.     

Selective inhibition of dipeptidyl peptidase 4 by targeting a substrate-specific secondary binding site

Dipeptidyl peptidase 4/CD26 (DP4) is a multifunctional serine protease liberating dipeptide from the N-terminus of (oligo)peptides which can modulate the activity of these peptides. The enzyme is involved in physiological processes such as blood glucose homeostasis and immune response. DP4 substrate specificity is characterized in detail using synthetic dipeptide derivatives. The specificity constant k(cat)/K(m) strongly depends on the amino acid in P?-position for proline, alanine, glycine and serine with 5.0 x 10? M?1 s?1, 1.8 x 10? M?1 s?1, 3.6 x 102 M?1 s?1, 1.1 x 102 M?1 s?1, respectively. By contrast, kinetic investigation of larger peptide substrates yields a different pattern. The specific activity of DP4 for neuropeptide Y (NPY) cleavage comprising a proline in P?-position is the same range as the k(cat)/K(m) values of NPY derivatives containing alanine or serine in P?-position with 4 x 10? M?1 s?1, 9.5 x 10? M?1 s?1 and 2.1 x 10? M?1 s?1, respectively. The proposed existence of an additional binding region outside the catalytic center is supported by measurements of peptide substrates with extended chain length. This 'secondary' binding site interaction depends on the amino acid sequence in P?'-P?'-position. Interactions with this binding site could be specifically blocked for substrates of the GRF/glucagon peptide family. By contrast, substrates not belonging to this peptide family and dipeptide derivative substrates that only bind to the catalytic center of DP4 were not inhibited. This more selective inhibition approach allows, for the first time, to distinguish between substrate families by substrate-discriminating inhibitors.     

A putative polyketide synthase/peptide synthetase from Magnaporthe grisea signals pathogen attack to resistant rice

Isolates of the rice blast fungus Magnaporthe grisea that carry the gene encoding Avirulence Conferring Enzyme1 (ACE1) are specifically recognized by rice (Oryza sativa) cultivars carrying the resistance gene Pi33. This recognition enables resistant plants to activate a defense response. ACE1 was isolated by map-based cloning and encodes a putative hybrid between a polyketide synthase and a nonribosomal peptide synthetase, enzymes involved in microbial secondary metabolism. ACE1 is expressed exclusively during fungal penetration of host leaves, the time point at which plant defense reactions are triggered. Ace1 appears to be localized in the cytoplasm of the appressorium. Mutation of the putative catalytic site of the beta-ketoacyl synthase domain of Ace1 abolishes recognition of the fungus by resistant rice. This suggests that Ace1 biosynthetic activity is required for avirulence. Our results are consistent with the hypothesis that the fungal signal recognized by resistant rice plants is the secondary metabolite whose synthesis depends on Ace1.     

云南地方猪种血液蛋白多态性研究

采用蛋白电泳技术研究了云南地方猪种血液蛋白多态性。共分析了3个云南地方猪种32-36个遗传位点,其中AKP、CAT、DIA、ES、G6PD、PA、6PGD、PHI、TF等9个 位点检测到多态性,多态位点百分比为0.1875-0.2121,平均杂合度为0.0712-0.1027。结果表明,云南地方猪种血液蛋白多态程度较高,反映在蛋白水平上的遗传多样性较为丰富。 Abstract:In this paper,protein electrophoresis was used to analyze the blood protein polymorphism in Yunnan local pig breeds and 32~36 genetic loci in the Yunnan local pig breeds were surveyed,Nine of them,such as AKP、CAT、DIA、ES、G6PD、PA、6PGD、PHI and TF,were found to be polymorphic,the mean heterozygosit(H)was 0.0712~0.1027.The results indicated that the blood protein polymorphism in the Yunnan local pig breeds is high,the Yunnan local pig breeds are wealthy in genetic diversity in point of their protein level.     

An expression profile of human α-lactalbumin in the milk of transgenic mouse

a-lactalbumin(a-Lac),amajorwheyprotein,isacalciummetalloprotein,thathasbeenfoundinallmilksstudiedsofar.ItinteractswithUDP-galactosyl-transferasetoformthelactosesynthetaseandthusmightbeakeyproteinforlactogenesis.Lactosesyn-thetaseispostulatedtobetherate-limitingenzymeforlactosebiosynthesis.Theincreaseda-Lacactivitycanproducesufficientlactosesynthetaseforthesynthesisoflactose,andinmilkyieldbydrawingwaterintomilk,sincelactoseisanosmoreactivemolecule.Transgenicswineoverexpressingbovinea-lactalbu…     

Multicolor chromosome banding (MCB) with YAC/BAC-based probes and region-specific microdissection DNA libraries

Multicolor chromosome banding (MCB) allows the delineation of chromosomal regions with a resolution of a few megabasepairs, i.e., slightly below the size of most visible chromosome bands. Based on the hybridization of overlapping region-specific probe libraries, chromosomal subregions are hybridized with probes that fluoresce in distinct wavelength intervals, so they can be assigned predefined pseudo-colors during the digital imaging and visualization process. The present study demonstrates how MCB patterns can be produced by region-specific microdissection derived (mcd) libraries as well as collections of yeast or bacterial artificial chromosomes (YACs and BACs, respectively). We compared the efficiency of an mcd library based approach with the hybridization of collections of locus-specific probes (LSP) for fluorescent banding of three rather differently sized human chromosomes, i.e., chromosomes 2, 13, and 22. The LSP sets were comprised of 107 probes specific for chromosome 2, 82 probes for chromosome 13, and 31 probes for chromosome 22. The results demonstrated a more homogeneous coverage of chromosomes and thus, more desirable banding patterns using the microdissection library-based MCB. This may be related to the observation that chromosomes are difficult to cover completely with YAC and/or BAC clones as single-color fluorescence in situ hybridization (FISH) experiments showed. Mcd libraries, on the other hand, provide high complexity probes that work well as region-specific paints, but do not readily allow positioning of breakpoints on genetic or physical maps as required for the positional cloning of genes. Thus, combinations of mcd libraries and locus-specific large insert DNA probes appear to be the most efficient tools for high-resolution cytogenetic analyses.     

早籼稻碾磨品质品种、地点、品种×地点互作效应的研究

以浙江省1995年早籼稻品种区域试验6个试点参试品种碾磨品质测定结果为试验材料,用多元分析法探讨了糙米率、精米率、整精米率3个碾磨品质的品种(基因型)、地点、品种×地点互作效应和各效应内碾磨品质间的相关关系,进行了参试品种互作效应检验。分析结果表明,糙米率、精米率以品种效应为主,整精米率性状以环境效应最大。相关分析显示,糙米率、精米率呈较强正相关,糙米率、精米率与整精米率均无相关关系。参试品种互作效应检验显示,多数品种整精米率均存在基因型×地点交互作用,糙米率、精米率多数品种较稳定。 Abstract:The genotype,test site,genotype×site effects on brown rice rate,milled rice recovery and head rice recovery as well as the correlation coefficients between the milling qualities within various effects were approached by using multiple analysis method with the data collected from early season indica rice varieties tested in regional trial in Zhejiang Province.Results showed that brown rice rate,milled rice recovery were predominantly affected by genotype.While head rice recovery was mostly conditioned by environment effect.Correlation analysis displayed that there were positive correlation between brown rice rate and milled rice rate.However,no correlation between brown rice rate,milled rice recovery and head rice recovery was found.The analysis of genotype×site of the varieties tested displayed that head rice recovery of the most tested varieties is involved with the interaction effects between the two components,while brown rice rate,milled rice recovery of the most tested varieties were more stable.     

大口鲇和鲇鱼血清蛋白质及同工酶的比较研究

采用聚丙烯酰胺梯度凝胶垂直板电泳,分析了大口鲇和鲇鱼的血清蛋白质以及心脏、肝脏、眼和肌肉4种组织的EST及MDH同工酶。结果表明,大口鲇和鲇鱼的血清蛋白质均能分离出20条左右的谱带,两者既表现出相同的谱带,又表现出迁移率和含量都不同的带型。两者的EST和MDH同工酶在4种组织及血清中均能特异性地表达,存在明显的组织和物种特异性。本文认为肝脏是研究大口鲇和鲇鱼种群生化遗传结构与变异的理想材料,同时还探讨了两种鲇鱼的M DH同工酶位点。 Abstract:The serum proteins and isozymes in four tissues (heart,liver,eye and musele)of Smeridionalis Chen and S.asotus Linnaeus were analyzed by polyacrylamide gradient gel vertical electrophoresis.The isozymes are esterase(EST)and malate dehytrogenase(MDH).The results showed that electrophoretograme of serum proteins were about 20 protein pattens in two species catfish,they were either the same protein pattens or the different pattens.Electrophoretogram of isozymes(EST,MDH)in two species catfish indicated tissues and species specificity.Experiment considered that the liver was a good material studied biochemical genetic constitution and variation in species group of S.meridionalis Chen and S.asotus Linnaeus.