月光花素甲(Ⅰ)、(Ⅱ)整体分子结构的测定

本文对月光花素甲(Calonyctin A)的整体分子结构分析加以完善并进行总结。前文报道月光花素甲是两个分子量相差28a.m.u的糖甙类同系物分子的混合物。这两个分子(简称M_1和M_2)分别称为月光花素甲(Ⅰ)与月光花素甲(Ⅱ)(Calonyctm A_1,CalonyctinA_2),分子量分别为938与910。它们分别含     

Capacity of Bacillus thuringiensis S-layer protein displaying polyhistidine peptides on the cell surface

S-layer protein of Bacillus thuringiensis strain CTC was used as the carrier protein to display polyhistidine (poly[6His]) peptides on the cell surface. Poly(6His) _n was fused with S-layer protein at two different sites, inserting just downstream of the S-layer protein homologous domain (slh) and replacing the non-slh region of S-layer protein, respectively. The two series chimeric proteins were both expressed by crystal negative B. thuringiensis strain 4Q7 and strain 171, respectively, as shown by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The recombinant B. thuringiensis cells gained Ni²⁺- and Cd²⁺-binding ability and had a capacity to display up to nine copies of poly(6His). The Cd²⁺ adsorption quantity of the recombinant strain with the strongest adsorption ability was twice that of the host strain.     

ClcR-based biosensing system in the detection of cis-dihydroxylated (chloro-)biphenyls

Polychlorinated biphenyls (PCBs) are a group of organic pollutants that are persistent when released into the environment. Among the metabolites of PCBs, dihydroxylated PCBs are also considered as toxic compounds. Various studies have shown that dihydroxylated PCBs affect the reproductive, immune, nervous, and endocrine systems. Detection of these chemicals in environmental and biological samples could provide first-hand information about their levels and lead to a better understanding of their role in toxicity. To that end, we developed a sensing system for the detection of dihydroxylated PCBs based on the clc operon. The Pseudomonas putida clc operon encodes a catabolic pathway for degradation of chlorocatechols, which are major metabolites of a large number of chlorinated compounds. In P. putida, the expression of these genes is regulated by a protein encoded by the gene clcR located upstream from the clcABD genes. We demonstrate here for the first time that dihydroxy PCBs can also induce the clc operon. Our sensing system employs P. putida bacteria harboring a plasmid in which the reporter gene, lacZ, is under the control of the regulatory protein ClcR. Consequently, when exposed to dihydroxy PCBs, the bacteria express β-galactosidase in an amount related to the concentration of the corresponding dihydroxy PCB. Various dihydroxylated PCBs, differing in the number and position of chlorines and in the position of hydroxyls, were tested for their ability to induce expression of β-galactosidase. Detection limits as low as 1×10⁻⁶ mol L⁻¹ were obtained for various dihydroxylated PCBs. Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

Heavy Vacuum Gas Oil Upregulates the Rhamnosyltransferases and Quorum Sensing Cascades of Rhamnolipids Biosynthesis in Pseudomonas sp. AK6U

We followed a comparative approach to investigate how heavy vacuum gas oil (HVGO) affects the expression of genes involved in biosurfactants biosynthesis and the composition of the rhamnolipid congeners in Pseudomonas sp. AK6U. HVGO stimulated biosurfactants production as indicated by the lower surface tension (26 mN/m) and higher yield (7.8 g/L) compared to a glucose culture (49.7 mN/m, 0.305 g/L). Quantitative real-time PCR showed that the biosurfactants production genes rhlA and rhlB were strongly upregulated in the HVGO culture during the early and late exponential growth phases. To the contrary, the rhamnose biosynthesis genes algC, rmlA and rmlC were downregulated in the HVGO culture. Genes of the quorum sensing systems which regulate biosurfactants biosynthesis exhibited a hierarchical expression profile. The lasI gene was strongly upregulated (20-fold) in the HVGO culture during the early log phase, whereas both rhlI and pqsE were upregulated during the late log phase. Rhamnolipid congener analysis using high-performance liquid chromatography-mass spectrometry revealed a much higher proportion (up to 69%) of the high-molecularweight homologue Rha–Rha–C₁₀–C₁₀ in the HVGO culture. The results shed light on the temporal and carbon source-mediated shifts in rhamonlipids’ composition and regulation of biosynthesis which can be potentially exploited to produce different rhamnolipid formulations tailored for specific applications.     

A new steroidal glycoside from Corypha taliera Roxb., a globally endangered species

The reversed-phased HPLC analysis of the methanol extract of the pericarp of C. taliera Roxb. (Talipalm), a rare species of Arecaceae family, afforded a new steroidal glycoside, β-sitosterol-3-O-α-l-rhamnopyranosyl-(1→4)-β-d-xylopyranosyl-(1→4)-β-d-glucopyranosyl-(1→4)-β-d-glucopyranoside (1). The structure of the compound was elucidated unequivocally by UV, IR, HR-ESI-MS, ¹H and ¹³C NMR spectroscopic studies.     

Design and synthesis of rhamnose-modified exenatide conjugate by sortase A-mediated ligation

Exenatide modified with rhamnose as a hapten was designed and synthesized by Sortase A-mediated ligation. An Exenatide peptide analog comprised of 46 amino acids, including the sortase A recognition motif LPETG at the C-terminus, was synthesized by solid phase peptide synthesis method. A tri-glycine modified-rhamnose derivative was chemically synthesized in seven steps in good yield. The site-specific conjugation between them catalyzed by Sortase A completed in 3?h and the rhamnose-modified Exenatide conjugate was obtained after semi-preparative HPLC purification and characterized by MALDI-TOF MS. This method should be generally useful for the synthesis of other Exenatide analog for biological studies.     

Composition, structure and solution properties of polysaccharides extracted from leaves of Mesembryanthenum crystallinum

Water and Chelator-soluble polymers were independently isolated from the alcoholic-insoluble substance (AIS) of Mesembryanthenum crystallinum leaves. After precipitation in ethanol and ultrafiltration (40 kD cut-off) of recovered solids dissolved in water, the yield relative of the so-call Lw and LCh polymers to the AIS dry matter was 2.5 ± 0.2 and 7 ± 0.5%. The galacturonic acid contents were 67 ± 3% and 63 ± 5%. The degrees of methylesterification of ca. 45 ± 3% and 50 ± 3%, showed that Lw and LCh belonged to the fairly methylesterified pectin class. From sugar analysis, LCh was shown to contain at least two types of pectic blocks, homogalacturonan (HG: 58%) and rhamnogalacturonan-I (RG-I: 34%). Their structure were deduced after saponification, polygalacturonase treatments, size exclusion chromatography (SEC) onto Sephacryl S-200 and then sugar composition of the collected fractions. The main polymers were methylesterified HG (32%), some of them being linked to non-methylesterified HG (10%), and to polygalacturonase-resistant HG (17%). In addition, there were (1) a highly soluble RG-I with long galactan side chains (RG-I-gal accounting for 9%) and (2) a RG-I with short arabino-galactan side chains (25%), named RG-I-ara/gal that was almost totally lost during SEC analysis, due to its low solubility at room temperature and in absence of chelators.SEC coupled with differential refractive index and light scattering showed that highly methylesterified HG chains exhibited aggregate structures in solution due to intermolecular hydrophobic interactions. These interactions formed hydrophobic clusters, which have been characterized by surface tension measurements and with a polarity probe, the Coomassie Brilliant Blue dye. After alkaline treatment of LCh, the self-assembly of HG disappeared.     

Spectral Study on the Interactions Among Cu(Ⅱ), Doxorubicin and CopC

Interactions among Cu(Ⅱ),doxorubicin and copper operon C(CopC)have been investigated in detail by means of fluorescence,UV-Vis,IR spectra,isothermal titration calorimetry(ITC)and molecular docking in Tris-HCl buffer(50 mmol/L,pH=7.4,25℃).The results suggest that Cu(Ⅱ)-doxorubicin is formed in a Cu(Ⅱ)to doxorubicin molar ratio of 1:2,and the conditional stability constant,K[Cu(Ⅱ)-doxorubicin]is 1.90×10^9 L^2/mol^2,CopC and doxorubicin can form a 1:1 complex,the conditional stability constant is greater than 10^5 L/mol.Binding of doxorabicin causes a conformational change in CopC with the reduction of β-sheet and increase of random coil,and the stability of CopC is decreased.Cu(Ⅱ),doxorubicin and CopC can form a CopC-Cu(Ⅱ)-doxorubicin ternary complex.The formation of CopC-Cu(Ⅱ)-doxorubicin reduced greatly the reduction rate of Cu(Ⅱ)by ascorbate(Vc),i.e.,the binding of doxorubicin affects the action of CopC as redox switch.     

单分子水平的酶催化与基因表达研究

近半个多世纪以来生命科学取得了非凡的进展, 从DNA双螺旋结构的提出, 到第一个酶晶体结构的被解析, 都得益于像X射线衍射、核磁共振、质谱这样的物理化学工具的发展. 如今, 在深入细致地定量研究生物活体系统中我们正面临新的挑战, 例如:了解酶及其他大分子复合物在体内是如何实时工作的, 它们在分子数很少时是怎样工作的, 在活细胞中大分子复合物是如何协调工作的, 以及不同的基因在活细胞中分子数很少的情况下是如何实现表达和不表达的等等. 近十多年来, 单分子成像, 超高分辨率显微镜和单分子操纵技术在世界范围内被广泛地运用于生物医学研究, 对生物化学和分子生物学的发展产生着深远的影响, 因为运用这些单分子、超高分辨技术, 使很多如上述的令人感兴趣的生物学问题实现了单分子层面上的研究和理解. 本文拟就近年来相关的物理化学方法特别是单分子方法和技术在生物医学中的应用做一个简要介绍.