Evolution of enzymatic activities in the enolase superfamily: galactarate dehydratase III from Agrobacterium tumefaciens C58.
Evolution of enzymatic activities in the enolase superfamily: galactarate dehydratase III from Agrobacterium tumefaciens C58.
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DOI：
10.1021/bi5005377
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发表时间：
2014-07-01
期刊：
影响因子：
2.9
通讯作者：
Gerlt, John A.
中科院分区：
文献类型：
--
作者：
Groninger-Poe, Fiona P.;Bouvier, Jason T.;Vetting, Matthew W.;Kalyanaraman, Chakrapani;Kumar, Ritesh;Almo, Steven C.;Jacobson, Matthew P.;Gerlt, John A.
根癌农杆菌C58的基因组编码烯醇化酶超家族（ENS）的12个成员，其中8个是扁桃酸消旋酶（MR）亚组的成员，因此，可能是酸性糖脱氢酶。使用77种酸性糖的文库进行高通量筛选，一种蛋白质（UniProt条目A9 CG 74;基因座标签Atu 4196）显示出与间半乳糖二酸和d-半乳糖醛酸的活性。以前在ENS中发现了两个半乳糖二酸酯酶家族，GalrD/TalrD [Yew，W.美国，等人（2007）Biochemistry 46，9564-9577]和GalrD-II [Rakus，J. F.，等人（2009）Biochemistry 48，11546-11558];这些具有不同的活性位点酸/碱催化，并且对d-半乳糖醛酸没有活性。A9 CG 74使间-半乳糖酸脱水形成2-酮-3-脱氧-半乳糖酸，但不像预期的那样使间-半乳糖醛酸脱水。相反，当A9 CG 74与d-半乳糖醛酸一起孵育时，形成3-脱氧-d-木糖-己二酸或3-脱氧-d-来苏糖-己二酸。在该反应中，不是将C5质子α提取到羧酸基团（ENS成员的预期反应），酶显然将质子α提取到醛基以形成3-脱氧-d-苏式-己糖醛酸，其经历类似于苄酸重排的1，2-氢化物移位以形成观察到的产物。A.根癌农杆菌C58在本研究中测试的条件下不利用间半乳糖二酸作为碳源，尽管它利用d-半乳糖醛酸，其可能是间半乳糖二酸的前体。编码A9 CG 74的基因和几个基因组近端基因用d-半乳糖醛酸作为碳源上调。其中之一是二氢吡啶二羧酸合酶超家族的成员，催化2-酮基-3-脱氧-d-半乳糖二酸脱水和随后脱羧为α-酮戊二酸半醛，从而提供了间-半乳糖二酸转化为α-酮戊二酸半醛的途径。
The genome of Agrobacterium tumefaciens C58 encodes 12 members of the enolase superfamily (ENS), eight of which are members of the mandelate racemase (MR) subgroup and, therefore, likely to be acid sugar dehydratases. Using a library of 77 acid sugars for high-throughput screening, one protein (UniProt entry A9CG74; locus tag Atu4196) showed activity with both m-galactarate and d-galacturonate. Two families of galactarate dehydratases had been discovered previously in the ENS, GalrD/TalrD [Yew, W. S., et al. (2007) Biochemistry46, 9564–9577] and GalrD-II [Rakus, J. F., et al. (2009) Biochemistry48, 11546–11558]; these have different active site acid/base catalysis and have no activity with d-galacturonate. A9CG74 dehydrates m-galactarate to form 2-keto-3-deoxy-galactarate but does not dehydrate d-galacturonate as expected. Instead, when A9CG74 is incubated with d-galacturonate, 3-deoxy-d-xylo-hexarate or 3-deoxy-d-lyxo-hexarate is formed. In this reaction, instead of abstracting the C5 proton α to the carboxylate group, the expected reaction for a member of the ENS, the enzyme apparently abstracts the proton α to the aldehyde group to form 3-deoxy-d-threo-hexulosuronate that undergoes a 1,2-hydride shift similar to the benzylic acid rearrangement to form the observed product. A. tumefaciens C58 does not utilize m-galactarate as a carbon source under the conditions tested in this study, although it does utilize d-galacturonate, which is a likely precursor to m-galactarate. The gene encoding A9CG74 and several genome proximal genes were upregulated with d-galacturonate as the carbon source. One of these, a member of the dihydrodipicolinate synthase superfamily, catalyzes the dehydration and subsequent decarboxylation of 2-keto-3-deoxy-d-galactarate to α-ketoglutarate semialdehyde, thereby providing a pathway for the conversion of m-galactarate to α-ketoglutarate semialdehyde.
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DOI：
10.1107/s0907444905036693
发表时间：
2006-01-01
影响因子：
2.2
作者：
Evans, P
通讯作者：
Evans, P
影响因子：
2.9
作者：
Bouvier JT;Groninger-Poe FP;Vetting M;Almo SC;Gerlt JA
通讯作者：
Gerlt JA
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
3.2
作者：
CHANG, YF;FEINGOLD, DS
通讯作者：
FEINGOLD, DS
影响因子：
48
作者：
Gibson, Daniel G.;Young, Lei;Smith, Hamilton O.
通讯作者：
Smith, Hamilton O.