Insights into microtubule nucleation from the crystal structure of human gamma-tubulin

Microtubules are hollow polymers of alphabeta-tubulin that show GTP-dependent assembly dynamics and comprise a critical part of the eukaryotic cytoskeleton. Initiation of new microtubules in vivo requires gamma-tubulin, organized as an oligomer within the 2.2-MDa gamma-tubulin ring complex (gamma-TuRC) of higher eukaryotes. Structural insight is lacking regarding gamma-tubulin, its oligomerization and how it promotes microtubule assembly. Here we report the 2.7-A crystal structure of human gamma-tubulin bound to GTP-gammaS (a non-hydrolysable GTP analogue). We observe a 'curved' conformation for gamma-tubulin-GTPgammaS, similar to that seen for GDP-bound, unpolymerized alphabeta-tubulin. Tubulins are thought to represent a distinct class of GTP-binding proteins, and conformational switching in gamma-tubulin might differ from the nucleotide-dependent switching of signalling GTPases. A crystal packing interaction replicates the lateral contacts between alpha- and beta-tubulins in the microtubule, and this association probably forms the basis for gamma-tubulin oligomerization within the gamma-TuRC. Laterally associated gamma-tubulins in the gamma-TuRC might promote microtubule nucleation by providing a template that enhances the intrinsically weak lateral interaction between alphabeta-tubulin heterodimers. Because they are dimeric, alphabeta-tubulins cannot form microtubule-like lateral associations in the curved conformation. The lateral array of gamma-tubulins we observe in the crystal reveals a unique functional property of a monomeric tubulin.     

Oxidative photochemical decarboxylation of zaragozic acid A

A unique decomposition reaction of the novel squalene synthase inhibitors called zaragozic acids has been studied. Under very mild conditions, e.g. by merely exposing their solutions to air and visible light at ambient temperature, these compounds, characterized by the 2,8-dioxabicyclo[3.2.1]octane-4,6,7-trihydroxy-3,4,5-tricarboxylic acid core, rapidly decompose. As relatively stable intermediates in the cascade of decomposition, the biologically active 2,8-dioxabicyclo[3.2.1]octane-6,7-dihydroxy-4-keto-5-caroxylic acid (or 3,4-decarboxy-4-dehydro) derivatives of these compounds have been isolated in ca. 20% yield. Derivatization on the highly reactive 4-carbonyl group yields stable derivatives, several of which are potent inhibitors of squalene synthase. Further decomposition results in the elimination of C3 and C4 atoms and the carboxylic acid on C5, the oxidation of C5 to carboxylic acid and the liberation of the oxo group on C1. Specific results obtained with zaragozic acid A, a key representative of the family of these potent cholesterol-lowering agents, are presented in this study.     

Mechanism limiting centrosome duplication to once per cell cycle

The centrosome organizes the microtubule cytoskeleton and consists of a pair of centrioles surrounded by pericentriolar material. Cells begin the cell cycle with a single centrosome, which duplicates once before mitosis. During duplication, new centrioles grow orthogonally to existing ones and remain engaged (tightly opposed) with those centrioles until late mitosis or early G1 phase, when they become disengaged. The relationship between centriole engagement/disengagement and centriole duplication potential is not understood, and the mechanisms that control these processes are not known. Here we show that centriole disengagement requires the protease separase at anaphase, and that this disengagement licences centriole duplication in the next cell cycle. We describe an in vitro system using Xenopus egg extract and purified centrioles in which both centriole disengagement and centriole growth occur. Centriole disengagement at anaphase is independent of mitotic exit and Cdk2/cyclin E activity, but requires the anaphase-promoting complex and separase. In contrast to disengagement, new centriole growth occurs in interphase, is dependent on Cdk2/cyclin E, and requires previously disengaged centrioles. This suggests that re-duplication of centrioles within a cell cycle is prevented by centriole engagement itself. We propose that the 'once-only' control of centrosome duplication is achieved by temporally separating licensing in anaphase from growth of new centrioles during S phase. The involvement of separase in both centriole disengagement and sister chromatid separation would prevent premature centriole disengagement before anaphase onset, which can lead to multipolar spindles and genomic instability.     

Ultracentrifugation of yeast cells in a gradient of acrylamide gel and sucrose

Zusammenfassung Dichtegefälle-Ultrazentrifugation, verbunden mit Photopolymerisation von Akrylamid, ist geeignet um lebende, unverletzte Zellen unbeweglich in Bänder zu trennen, insofern sie sich punkto Gewicht, Volumen, Dichte oder Beweglichkeit im Dichtegefälle unterscheiden. Verschieden ernährte Hefezellen-Gruppen (Candida albicans) zeigten verschiedene Bandenverteilungen.     

承载供应链管理的客户关系链管理分析与建模

指出了客户关系管理的概念不足,以实际例子引出客户关系链的概念,并分析指出客户关系链可以承载供应链的运作管理.利用Markov链对该过程建模,对简单的情况进行了分析和计算,并给出了如果忽略了客户下游信息,企业所受到的损失分析.并同供应链系统作了类比.     

一个用于中小型企业的扩展ERP软件流程集成模型分析

在以前工作的基础上,主要讨论了一个将ERP(enterpriseresourceplanning,企业资源规划)、SRM(supplierrelationshipmanagement,供应商关系管理)、CRM(customerrelationshipmanagement,客户关系管理)集成为一体的简化流程模型.论文在界定了几个基本概念的基础上,描述了接触中心的发出流程和接收流程功能,讨论了对ERP软件的流程及核心管理和CRM软件的流程及核心管理思路,研究了对ERP同CRM的关联点,仿照对CRM的分析,对SRM做了类似的分析,最后给出了以关系为企业围墙的扩展ERP简化模型.