Glycogen debranching enzyme: purification, antibody characterization, and immunoblot analyses of type III glycogen storage disease.

Type III glycogen storage disease is caused by a deficiency of glycogen debranching-enzyme activity. Many patients with this disease have both liver and muscle involvement, whereas others have only liver involvement without clinical or laboratory evidence of myopathy. To improve our understanding of the molecular basis of the disease, debranching enzyme was purified 238-fold from porcine skeletal muscle. In sodium dodecyl sulfate-polyacrylamide gel electrophoresis the purified enzyme gave a single band with a relative molecular weight of 160,000 that migrated to the same position as purified rabbit-muscle debranching enzyme. Antiserum against porcine debranching enzyme was prepared in rabbit. The antiserum reacted against porcine debranching enzyme with a single precipitin line and demonstrated a reaction having complete identity to those of both the enzyme present in crude muscle and the enzyme present in liver extracts. Incubation of antiserum with purified porcine debranching enzyme inhibited almost all enzyme activity, whereas such treatment with preimmune serum had little effect. The antiserum also inhibited debranching-enzyme activity in crude liver extracts from both pigs and humans to the same extent as was observed in muscle. Immunoblot analysis probed with anti-porcine-muscle debranching-enzyme antiserum showed that the antiserum can detect debranching enzyme in both human muscle and human liver. The bands detected in human samples by the antiserum were the same size as the one detected in porcine muscle. Five patients with Type III and six patients with other types of glycogen storage disease were subjected to immunoblot analysis. Although anti-porcine antiserum detected specific bands in all liver and muscle samples from patients with other types of glycogen storage disease (Types I, II, and IX), the antiserum detected no cross-reactive material in any of the liver or muscle samples from patients with Type III glycogen storage disease. These data indicate (1) immunochemical similarity of debranching enzyme in liver and muscle and (2) that deficiency of debranching-enzyme activity in Type III glycogen storage disease is due to absence of debrancher protein in the patients that we studied.     

Simultaneous convective and diffusive mass transfers in a hemodialyser

The mass transfer in a hemodialyser in the presence of combined dialysis and ultrafiltration has been calculated by integration of mass fluxes across the boundary layers in blood and dialysate phase taking into account the partial rejection of solute as well as changes in local blood flow and ultrafiltration flux along the membrane. Clearances of creatinin, vitamin B12, and myoglobin have been calculated as a function of blood and ultrafiltrate flow rate and were found to be in good agreement with in vitro measurements. The data suggest the following empirical correlation for the hemodiafiltration clearance.     

盐酸胍诱导的α淀粉酶去折叠与再折叠

利用蛋白质内源荧光和酶活性两种信号以及荧光偏振,HPLC和停流等方法研究了盐酸胍诱导的α淀粉酶去折叠与重折叠的平衡转变和动力学。实验结果表明α淀粉酶去折叠与重折叠是两个不同的过程;变性与复性过程中可能伴有聚集体生成;去折叠与重折叠均为双相过程,重折叠大约始于2秒之后。     

蛋白质结构成对比较的新方法

介绍一种蛋白质三维结构的快速比较方法.此方法毋需初始联配,而能自动寻找和智能迭代.利用本程序对珠蛋白、丝氨酸蛋白酶、天冬氨酸蛋白酶、钙结合蛋白和溶菌酶作了系统的结构比较,取得了满意的结果.本文也讨论了衡量联配结果好坏的要素问题.     

中药固真方对一些与细胞增殖有关基因表达的影响

 中药固真方对一些与细胞增殖有关基因表达的影响姚明忠,顾文聪,丁卫,韩志芬,杜国光（上海中医药大学生物化学教研室,上海２０００３２）（北京医科大学生物化学与分子生物学系,北京１０００８３）中药固真方（ＶＲＦ）具有补肾益精、延缓衰老的作用［１］．能提高成...     

有机盐对基因启动子的σ因子选择性的影响

用σ￣（７０）或σ￣（３８）和核心酶（Ｅ）组成的大肠杆菌ＲＮＡ聚合酶全酶（Ｅσ）对四种启动子进行了体外转录。结果表明：ｌａｃＵＶ５,ｒｐｌＪ主要被Ｅσ￣（７０）识别,ｋａｔＥ主要被Ｅσ￣（３８）识别,ｆｉｃ在低盐浓度时被Ｅσ￣（７０）识别,在高浓度盐时被Ｅσ￣（３８）识别;Ｅσ￣（３８）转录特异性启动子所需酶量大于Ｅσ￣（７０）转录特异性启动子的酶量;在含有分别对σ￣（７０）或σ￣（３８）亲和性大的混和启动子的体外转录中,启动子之间不存在干扰和竞争,转录水平与启动子浓度成正相关。     

Mass spectra of underivatized peptide amides related to substance P

Mass spectra of underivatized hexa- and heptapeptide amides related to Substance P have been obtained with a conventional electron ionization mass spectrometer using sample vaporization from a tungsten wire by the technique of rapid heating, proton transfer ionization using ammonia, and photoplate recording of spectra. These spectra exhibit little evidence of sample pyrolysis and are readily interpreted to yield amino acid sequences.     

Prostaglandin production in the umbilical and uterine circulations in pregnant sheep at 129-136 days gestation.

Prostaglandins circulating in the maternal and foetal blood have been implicated in important physiological systems. These functions include foetal adrenal function, maintenance of patency of the ductus arteriosus, regulation of uterine and umbilical circulations, and labor and delivery type myometrial contractions. The placenta is a major site of prostaglandin production in pregnancy. Limited data are available which combine measurements of veno-arterial differences across the uterine and umbilical circulations with blood flow in these circulations to enable calculation of umbilical-placental and utero-placental production rates for the prostaglandins. In chronically instrumented pregnant ewes, between 129 and 136 days of gestation, prostaglandin F2 alpha(PGF2 alpha), 13, 14 dihydro-15-keto prostaglandin F2 alpha (PGFM), prostaglandin E2 (PGE2) were measured in the maternal carotid artery and uterine vein. Foetal PGE2, and 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha) (the major metabolite of prostacyclin) were measured in umbilical venous and foetal descending aorta arterial plasma. Umbilical and uterine blood flow were measured using the diffusion-equilibrium technique. Uterine blood flow was 1693 +/- 137 ml.min-1 (mean +/- SEM); uterine production rates were 480 +/- 88 ng.min-1 for PGF2 alpha, 517 +/- 144 ng.min-1 for PGFM, and 165 +/- 27 ng.min-1 for PGE2. Umbilical blood flow was 147 +/- 17 ml.min-1.kg-1 foetal body weight. Umbilical production rates into the foetal circulation were 11 +/- 2 ng.min-1.kg-1 for PGE2 and 6 +/- 2 ng. ng.min-1.kg-1 foetal body weight for PGI2.     

Development of mechanistically based model for simulating soluble microbial products generation in an aerated/non-aerated SBR

Soluble microbial products (SMPs) are considered as the main organic components in wastewater treatment plant effluent from biological wastewater treatment systems. To investigate and explore SMP metabolism pathway for further treatment and control, two innovative mechanistically based activated sludge models were developed by extension of activated sludge model no.3 (ASM3). One was the model by combining SMP formation and degradation (ASM3-SMP model) processes with ASM3, and the other by combining both SMP and simultaneous substrate storage and growth (SSSG) mechanisms with ASM3 (SSSG-ASM3-SMP model). The detailed schematic modification and process supplements were introduced for comprehensively understanding all the mechanisms involved in the activated sludge process. The evaluations of these two models were demonstrated by a laboratory-scale sequencing batch reactor (SBR) operated under aerated/non-aerated conditions. The simulated and measured results indicated that SMP comprised about 83% of total soluble chemical oxygen demand (SCOD) in which biomass-associated products (BAPs) were predominant compared with utilization-associated products (UAPs). It also elucidated that there should be a minimum SMP value as the reactive time increases continuously and this conclusion could be used to optimize effluent SCOD in activated sludge processes. The comparative results among ASM3, ASM3-SMP and SSSG-ASM3-SMP models and the experimental measurements (SCOD, ammonia and nitrate nitrogen) showed clearly the best agreement with SSSG-ASM3-SMP simulation values (R = 0.993), strongly suggesting that both SMP formation and degradation and SSSG mechanisms are necessary in biologically activated sludge modeling for municipal wastewater treatment.     

DNA methylation changes in photoperiod-thermo-sensitive male sterile rice PA64S under two different conditions

Epigenetic modification can occur at a high frequency in crop plants and might generate phenotypic variation without changes in DNA sequences. DNA methylation is an important epigenetic modification that may contribute to environmentally-induced phenotypic variations by regulating gene expression. Rice Photoperiod-Thermo-Sensitive Genic Male Sterile (PTGMS) lines can transform from sterility to fertility under lower temperatures and short-day (SD) conditions during anther development. So far, little is known about the DNA methylation variation of PTGMS throughout the genome in rice. In this study, we investigated DNA cytosine methylation alterations in the young panicles of PTGMS line PA64S under two different conditions using methylation sensitive amplified polymorphism (MSAP) method. Compared with the DNA methylation level of PA64S under lower temperatures and SD conditions (fertility), higher methylation was observed in PA64S (sterility). The sequences of 25 differentially amplified fragments were successfully obtained and annotated. Three methylated fragments, which are homologous to D2, NAD7 and psaA, were confirmed by bisulfite sequencing and their expression levels were also evaluated by qPCR. Real time quantitative PCR analysis revealed that five of the six selected methylated genes were downregulated in PA64S (sterility). These results suggested that DNA methylation may be involved in the sterility–fertility transition of PA64S under two different environmental conditions.