Biochemical characterization and crystallization of porin from Rhodopseudomonas blastica

Oligomeric porin of the phototrophic bacterium Rhodopseudomonas blastica DSM 2131 was obtained from cell envelopes by differential temperature extraction in the presence of detergent and salt. The isolated porin exhibited strong porin activity after reconstitution into lipid bilayer membranes. The effective channel diameter for the trimer was estimated as 1.5 nm from single channel conductance measurements in the presence of 1 M KCl. Moderate cation-selectivity was observed. Oligomeric porin migrated as a single band (apparent molecular weight 81 kDa) on sodium dodecyl sulfate polyacrylamide gelelectrophoresis when solubilized below 70 °C. The oligomers were converted into monomers on heating to 70 °C or above forming two bands with apparent molecular weight of 36 kDa and 35 kDa. The oligomer was not sensitive to EDTA. Its molecular weight was determined to be 119.3 kDa by analytical ultracentrifugation. The isoelectric point was 5.7. Circular dichroism data indicated a high content of -sheet structure. Gasphase sequencing of the N-terminal residues revealed the sequence: NH₂-Glu-Ile-Ser-Leu-Asn-Gly-Tyr-Gly-Arg-Phe. Crystals with a maximal side length of 300 m and diffracting to 0.32 nm resolution were obtained with the porin oligomer in the presence of C8E4 and 1,2,3-heptanetriol by using the vapor phase equilibration technique.Abbreviations C8E4 n-octyl tetraoxyethylene - M_r apparent molecular weight - Octyl-POE n-octyl polyoxyethylene - LDAO N,N-dimethyl dodecyl aminoxide - LPS lipopolysaccharide - PAGE polyacrylamide gel-electrophoresis - PEG polyethylene glycol     

Methyl chloride metabolism of the strictly anaerobic,methyl chloride-utilizing homoacetogen strain MC

The methyl chloride metabolism of the homoacetogenic, methyl chloride-utilizing strain MC was investigated with cell extracts and cell suspensions of the organism. Cell extracts were found to contain all enzyme activities required for the conversion of methyl chloride or of H₂ plus CO₂ to acetate. They catalyzed the dechlorination of methyl chloride with tetrahydrofolate as the methyl acceptor at a rate of 20 nmol/min × mg of cell protein. Also, the O-demethylation of vanillate with tetrahydrofolate could be measured at a rate of 40 nmol/min × mg. Different enzyme systems appeared to be responsible for the dehalogenation of CH₃Cl and for the O-demethylation of methoxylated aromatic compounds, since cells grown with methoxylated aromatic compounds exhibited a significantly lower activity of CH₃Cl conversion than methyl chloride grown cells and vice versa. In addition, ammonium thiocyanate (5 mM) completely inhibited CH₃Cl dechlorination, whereas the consumption of vanillate was not affected significantly. The data were taken to indicate, that the methyl chloride dehalogenation is catalyzed by a specific, inducible enzyme present in strain MC, and that tetrahydrofolate rather than the corrinoid-protein involved in acetate formation is the primary acceptor of the methyl group in the dechlorination reaction.     

Formation of acetyl-CoA in Zymomonas mobilis by a pyruvate dehydrogenase complex

In the gram negative, obligately ethanologenic bacterium Zymomonas mobilis a pyruvate dehydrogenase complex was identified and the complex was enriched from cell extracts. This multienzyme complex is responsible for acetyl-CoA biosynthesis from pyruvate. No activities of related multienzyme complexes, 2-ketoglutarate dehydrogenase and branched chain keto acid dehydrogenase, could be detected.     

Rates of decay for the survival probability of a mutant gene II The multitype case

This paper extends the results of [1] to the multitype case. For a multitype branching process that is slightly supercritical, approximations for the survival probability in terms of the maximal eigenvalue of the mean matrix and a generalized variance ² are developed. Our results improve upon those of Hoppe [5] and Eshel [3] that seek to validate a conjecture of Ewens [4].Research supported in part by NSF grant DMS 9007182     

多聚半乳糖醛酸内切酶的固定化及其性质研究

棉花枯萎病菌多聚半乳糖醛酸内切酶在pH大于7时不稳定,故对它进行多种化学修饰而又不影响其活性,必须在pHd小于7的体系中进行。本文报道将PGAUase在还原剂存在下,与稀酸处理的Sepharose 4B交联,获得较高活力的固定化酶。固定化酶催化动力学表明,最适pH为4,4,最适温度为55℃,在pH1至8.0范围内稳定。和溶液酶比较,对热稳定性提高,但对碱稳定性下降。以多聚半乳糖醛酸为底物,Km为0.27mmol/L,Vmax为66.67nmol/L·min,均大于溶液酶(Km=0.07mmol/L,Vmax=28.00nmol/L·min)。在pH4.8,30℃,聚半乳糖醛酸在固相酶的柱中循环水解不同的时间降解产物经圆盘电泳和等电聚焦测定,得到不同大小的寡糖片段混合物,证明固相酶和溶液酶的作用方式相同,同时使以酶解法制备一定大小的有生物活性的寡糖分子成为可能。     

大鼠肾脏和其它组织中磷酸酪氨酸蛋白的研究

本文采用P-tyr-BSA为免疫原免疫家无得抗血清。将纯化的IgG与HRP偶联,建立了P-tyr-Pr的ELISA法,并测定了正常大鼠肾脏等组织中P-tyr-Pr含量,其分布规律如下:上清中P-tyr-Pr含量高者,其颗粒部分则低,反之亦然;其中肾脏上清中含量远比其它组织(脾、肺、肝等)高。在此基础上,又研究了膜性肾炎大鼠肾脏P-tyr-Pr含量,发现其上清中的含量远远高于正常大鼠肾脏中的含量。     

Hydrolysis of Butteroil by Immobilized Lipase Using a Hollow-Fiber Reactor: Part VI. Multiresponse Analyses of Temperature and pH Effects

A lipase from Aspergillus niger, immobilized by physical adsorption on hydrophobic hollow fibers made of microporous polypropylene, was used to effect the hydrolysis of the glycerides of melted butterfat at 40, 50, 55, and 60°C (pH 7.0), and at pH 3.0, 4.0, 5.0, 7.0, 8.0, and 9.0 (40°C). McIlvane buffer and melted butterfat were pumped cocurrently through the hollow fiber reactor. The concentrations of ten different free fatty acids in the effluent oil stream were measured by HPLC. Multiresponse nonlinear regression methods were employed to fit the data to multisubstrate rate expressions derived from a Ping Pong Bi Bi mechanism in which the rate controlling step is deacylation of the enzyme. Thermal deactivation of the immobilized lipase was also included in the mathematical model of reactor performance. A postulated normal distribution of v_max with respect to the number of carbon atoms of the fatty acid residue (with an additive correction for the number of double bonds) was found to provide the best statistical fit of the data. The models developed can be used to independently predict the effects of either the pH or the temperature, as well as the reactor space time and the time elapsed after immobilization, on the free fatty acid profile of the lipolyzed butteroil product.     

Multiple origins of self-fertilization in tristylous Eichhornia paniculata (Pontederiaceae): Inferences from style morph and isozyme variation

Populations of Eichhornia paniculata (Pontederiaceae) exhibit a wide range of mating systems, from predominant outcrossing to high levels of self-fertilization. The origin of self-fertilization in this tristylous species is associated with the loss of style-length morphs from populations and the spread of self-pollinating, floral variants. We examined geographic variation in style morph and allozyme frequencies to determine whether the loss of style morphs and transition to selfing could have multiple origins in E. paniculata. Surveys of floral variation in 167 populations from six states in northeastern Brazil revealed that at least one style morph was absent from 29.3%. Non-trimorphic populations occurred in all states and ranged in frequency from 9% in Ceará to 68% in Alagoas. Selfing variants occurred in 8.5% and 55% of trimorphic and non-trimorphic populations, respectively, and were distributed among five of six states with primary concentrations in Alagoas and Pernambuco. A comparison of electrophoretic variation at 24 isozyme loci in 28 trimorphic, 13 dimorphic and 3 monomorphic populations indicated that non-trimorphic populations contained 84% of the allelic variation present in trimorphic populations and were markedly differentiated from one another. Analyses of genetic distance and the distribution of rare alleles indicated that non-trimorphic populations were often more similar to neighbouring trimorphic populations than to one another. Populations with selfing variants occurred at low frequency in three genetically distinct parts of the range. These results, in combination with genetic and morphological evidence suggest that style morphs are lost repeatedly from populations of E. paniculata and that selfing variants may have originated on at least three separate occasions in northeastern Brazil.     

Are brood sex ratios adaptive?—The effect of experimentally altered brood sex ratio on nestling growth,mortality and recruitment

Brood sex ratios (BSRs) have often been found to be nonrandom in respect of parental and environmental quality, and many hypotheses suggest that nonrandom sex ratios can be adaptive. To specifically test the adaptive value of biased BSRs, it is crucial to disentangle the consequences of BSR and maternal effects. In multiparous species, this requires cross-fostering experiments where foster parents rear offspring originating from multiple broods, and where the interactive effect of original and manipulated BSR on fitness components is tested. To our knowledge, our study on collared flycatchers (Ficedula albicollis) is the first that meets these requirements. In this species, where BSRs had previously been shown to be related to parental characteristics, we altered the original BSR of the parents shortly after hatching by cross-fostering nestlings among trios of broods and examined the effects on growth, mortality and recruitment of the nestlings. We found that original and experimental BSR, as well as the interaction of the two, were unrelated to the fitness components considered. Nestling growth was related only to background variables, namely brood size and hatching rank. Nestling mortality was related only to hatching asynchrony. Our results therefore do not support that the observed BSRs are adaptive in our study population. However, we cannot exclude the possibility of direct effects of experimentally altered BSRs on parental fitness, which should be evaluated in the future. In addition, studies similar to ours are required on various species to get a clearer picture of the adaptive value of nonrandom BSRs.     

我国大麦和性花叶病毒（BaMMV）研究初报

本文根据F(ab′)2－ELISA和ISEM实验,结合病毒粒子长度测定,报道了江苏省如东县农科所大麦黄花病毒源（吕种盐辐矮早3）中除存在大麦花花叶病毒（BaYMV）外,还复合感染大麦和性花叶病毒（BaMMV）,这是我国BaMMV的第一次报道。