The bacterial class IIIb adenylate cyclase CyaC-Sm of Sinorhizobium meliloti: Redox sensing by a heme B binding membrane anchor

原核和真核腺苷酸环化酶：具有感觉功能的膜锚？腺苷酸环化酶（AC）从ATP产生第二信使环AMP存在于所有种类的生命（真核生物、细菌和古细菌）中。cAMP在调节细胞功能方面具有广泛的意义。AC酶被分为六类，并且最好表征的AC是涵盖大多数真核生物和细菌AC的III类的成员。这类AC在结构域组成、信号感知、调节和整体功能方面表现出很大的变化。本研究旨在了解苜蓿中华根瘤菌CyaC-Sm的功能及其六螺旋（6-TM）膜锚在氧化还原传感中的作用。6-TM膜锚在真核生物（包括哺乳动物）和一些细菌III类AC中是保守的。这种设计的细菌AC尚未在分子细节上进行表征。了解CyaC-Sm可能对评估膜锚及其在真核III类AC中的潜在感觉作用产生影响。准备工作表明，CyaC-Sm含有醌反应血红素B在膜锚。呼吸和醌应该氧化铁氧还蛋白型结构域中的血红素B和铁硫簇，从而控制催化结构域的二聚化和活性。膜锚与醌/血红素网站和附近的FeS簇显示出显着的相似性，以及其特征在于呼吸系统延胡索酸还原酶的结构和功能方面的域。将用纯化的CyaC-Sm和在大肠杆菌中表达的野生型和突变型CyaC-Sm来表征组分和连接的氧化还原酶（SMc 01816）在控制AC活性中的作用。杆菌总的来说，这项研究的目的是在功能和CyaC-Sm作为一种新的，实际上是第一个，氧化还原控制的III类腺苷酸环化酶的分子细节的特征。这项工作可能具有更广泛的意义，通过分配AC蛋白的六螺旋TM结构域的感觉作用，也讨论了膜界定的真核AC。CyaC-Sm在S.建议书中没有分析苜蓿草。然而，有第一个迹象表明，CyaC-Sm是连接到有氧呼吸和反硝化作用。

Pro- and eukaryotic adenylate cyclases: Membrane anchors with sensory function?Adenylate cyclases (AC) producing the second messenger cyclic AMP from ATP are present in all classes of life (eukarya, bacteria and archaea). Cyclic AMP has a broad significance in regulating cellular functions. The AC enzymes are grouped in six classes, and the best characterized ACs are members of class III which covers most eukaryotic and bacterial ACs. The ACs of this class show a large variation in domain composition, signal perception, regulation and overall function. The proposal aims at understanding the function of CyaC-Sm of Sinorhizobium meliloti, a class IIIb AC, and the role of its hexa-helical (6-TM) membrane anchor in redox sensing. A 6-TM membrane anchor is conserved in the eukaryotic (including mammalian) and some bacterial class III ACs. Bacterial ACs of this design have not been characterized in molecular detail. Understanding CyaC-Sm may have an impact on assessing the membrane anchor and its potential sensory role in eukaryotic class III ACs. Preparatory work suggests that CyaC-Sm contains a quinone reactive heme B in the membrane anchor. Respiration and quinones are supposed to oxidize the heme B and an iron-sulfur cluster in a ferredoxin-type domain and, as a result, to control dimerization and activity of the catalytic domain. The membrane anchor with the quinone/heme site and the near-by FeS cluster show a remarkable similarity to domains of the well-characterized respiratory fumarate reductase in structural and functional terms. The role of the components and of a linked oxidoreductase (SMc01816) in controlling AC activity will be charac-terized with purified CyaC-Sm, and by the use of wild-type and mutant CyaC-Sm expressed in E. coli. Overall, the study aims at characterizing function and molecular details of CyaC-Sm as a novel, actually the first, redox-controlled class III adenylate cyclase. The work might be of broader significance by assigning the hexa-helical TM domain of AC proteins a sensory role which is also discussed for membrane-delimited eukaryotic ACs. The physiological role of CyaC-Sm in S. meliloti is not analyzed in the proposal. However, there are first indications that CyaC-Sm is linked to aerobic respiration and to denitrification.