Production, crystallization, and preliminary X-ray analysis of rabbit skeletal muscle troponin complex consisting of troponin C and fragment (1-47) of troponin I.

Troponin is a ternary protein complex consisting of subunits TnC. TnI, and TnT, and plays a key role in calcium regulation of the skeletal and cardiac muscle contraction. In the present study, a partial complex (CI47) was prepared from Escherichia coli-expressed rabbit skeletal muscle TnC and fragment 1-47 of TnI, which is obtained by chemical cleavage of an E. coli-expressed mutant of rabbit skeletal muscle TnI. Within the ternary troponin complex, CI47 is thought to form a core that is resistant to proteolytic digestion, and the interaction within CI47 likely maintains the integrity of the troponin complex. Complex CI47 was crystallized in the presence of sodium citrate. The addition of trehalose improved the diffraction pattern of the crystals substantially. The crystal lattice belongs to the space group P3(1)(2)21, with unit cell dimensions a = b = 48.2 A, c = 162 A. The asymmetric unit presumably contains one CI47 complex. Soaking with p-chloromercuribenzenesulfonate (PCMBS) resulted in loss of isomorphism, but enhanced the quality of the crystals. The crystals diffracted up to 2.3 A resolution, with completeness of 91% and R(merge) = 6.4%. The crystals of PCMBS-derivative should be suitable for X-ray studies using the multiple-wavelength anomalous diffraction technique. This is the first step for elucidating the structure of the full troponin complex.     

Two independent growth factor-generated signals regulate c-fos and c-myc mRNA levels in Swiss 3T3 cells

Polypeptide growth factors that stimulate cell proliferation bind to cell surface receptors and activate intracellular signal transduction pathways. One major signalling pathway, initiated by phosphatidylinositol (PI) turnover, involves activation of protein kinase C. Some polypeptide growth factors, including mitogens that activate protein kinase C, induce a rapid increase in expression of the proto-oncogenes, c-myc and c-fos. In order to characterize the signal transduction pathways responsible for proto-oncogene activation, we treated Swiss 3T3 cells with the tumor promoter phorbol dibutyrate to generate cells deficient in protein kinase C. These cells were then stimulated with platelet extract, bombesin, or epidermal growth factor (EGF) and the levels of c-myc and c-fos mRNA were determined. Platelet extract or bombesin, which stimulate PI turnover, were substantially weaker inducers of c-myc and c-fos mRNA levels in the protein kinase C-depleted cells, although some variability with platelet extract was noted. EGF, which does not stimulate PI turnover in several cell systems, was by contrast a potent inducer of both proto-oncogenes whether or not the cells were deficient in protein kinase C. Pretreatment of cells with phorbol dibutyrate caused little or no change in the basal levels of c-myc or c-fos mRNA, but led to a small but significant increase in basal levels of ornithine decarboxylase mRNA. These results demonstrate that EGF and growth factors that activate PI turnover induce expression of the c-myc and c-fos proto-oncogenes through different pathways.     

Direct micro-sequence analysis of peptides from Escherichia coli ribosomal proteins S11, L9 and L29 after separation by reversed phase chromatography

Tryptic peptides of the ribosomal proteins S11, L9 and L29 were separated by reversed phase chromatography under conditions which enabled direct micro-sequencing with the 4-(dimethylamino)azobenzene-4'-isothiocyanate/phenylisothiocyanate double coupling method [Chang, Brauer, Wittmann-Liebold (1978) FEBS Lett. 93, 205-214]. The peptides were separated on a RP-18 column employing volatile buffers at pH 2.0, 4.1 and 7.8. Depending on the different chromatographic behaviour of the peptide mixture, the elution gradient was optimised for each hydrolysate using 20 micrograms of the hydrolysed protein. Preparative separations were made with 150-250 micrograms. At least 80% of the peptides could be isolated by these techniques and used for direct micro-sequencing without further purification or desalting. The results show that the high-performance liquid chromatographic method employed allows easy isolation and sequencing with minute amounts of peptides.     

Conversion of sterigmatocystin to aflatoxin B 1 by Aspergillus parasiticus

¹⁴C-Sterigmatocystin isolated from cultures of $\text{Aspergillus}$$\text{versicolor}$ supplemented with (1-¹⁴C)acetate was shown to be efficiently converted to aflatoxin B₁ by the resting mycelium of $\text{A. parasiticus}$. The experimental results may indicate a biosynthetic pathway leading from 5-hydroxysterigmatocystin to sterigmatocystin and then to aflatoxin B₁.     

Research Progress on Anti-Inflammatory Mechanisms of Black Ginseng

In recent years, black ginseng, a new type of processed ginseng product, has attracted the attention of scholars globally. Ginsenoside and ginseng polysaccharide, the main active substances of black ginseng, have been shown to carry curative effects for many diseases. This article focuses on the mechanism of their action in anti-inflammatory response, which is mainly divided into three aspects: activation of immune cells to exert immune regulatory response; participation in inflammatory response-related pathways and regulation of the expression level of inflammatory factors; effect on the metabolic activity of intestinal flora. This study identifies active anti-inflammatory components and an action mechanism of black ginseng showing multi-component, multi-target, and multi-channel characteristics, providing ideas and a basis for a follow-up in-depth study of its specific mechanism.     

Ce~（3＋）与G－肌动蛋白结合引起的构象变化及对聚合的影响

用Ａｅｄａｎｓ标记肌动蛋白单体Ｇ－Ａｃｔｉｎ上Ｃｙｓ３７４残基作为探针,研究了稀土离子Ｃｅ~（３＋）与Ｇ－Ａｃｔｉｎ的结合及引起的微构象变化。Ｃｅ~（３＋）在低浓度（Ｃｅ~（３＋）／Ａｃｔｉｎ摩尔比＜１）和Ｃａ~（２＋）竞争Ｇ－Ａｃｔｉｎ上二价离子的高亲合位点。Ｃｅ~（３＋）取代Ｃａ~（２＋）引起Ａｅｄａｎｓ荧光强度增强与Ｍｇ~（２＋）取代Ｃａ~（２＋）的结果相同。Ｃｅ~（３＋）／Ａｃｔｉｎ＞ｌ则导致Ａｅｄａｎｓ荧光强度下降。说明Ｃｅ~（３＋）在高低两种浓度条件下结合的位点及对Ｃｖｓ３７４的微构象的影响不同。时间分辩测得的Ａｅｄａｎｓ荧光寿命也支持这一结论。ＣＤ谱结果表明Ｃｅ~（３＋）／Ａｃｔｉｎ＜０．４,Ａｃｔｉｎ的二级结构增加,大于０．４又导致其失去。Ｃｅ~（３＋）－Ａｃｔｉｎ在有／无游离ＡＴＰ时用聚合液诱导的聚合结果表明,无游离ＡＴＰ时,极低浓度Ｃｅ~（３＋）促进聚合,高浓度虽有促进但有所减弱;有游离ＡＴＰ时,Ｃｅ~（３＋）／Ａｃｔｉｎ在实验范围内促进聚合。     

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...     

Serum lipoproteins and albumin in the lecithin: Cholesterol acyltransferase reaction

The unique features of pig ovarian follicular fluids, i.e., presence of high density lipoprotein (HDL) only and lecithin: cholesterol acyltransferase (EC 2.3.1.43; LCAT) activity, provides a good model to study the effect of serum lipoproteins and serum albumin on the LCAT reaction. $\text{In}\text{vitro}$ cholesterol esterification is enhanced when very low density lipoprotein (VLDL) and low density lipoprotein (LDL) fractions are added, but is inhibited when one or the other of these lipoproteins is absent. High concentrations of HDL₂ result in decreased activation which can be compensated for by the addition of the VLDL-LDL mixture. These findings suggest that the rate of cholesterol esterification in ovarian follicular fluid may be enhanced by providing the exogenous VLDL and LDL as the recipients of HDL-cholesteryl ester. The inhibition of LCAT activity caused by free fatty acid and lysophosphatidylcholine can be partially reversed by the addition of serum albumin, suggesting that serum albumin may regulate the LCAT reaction.