您的位置 首页 生物催化剂

ACS Catalysis：具备多底物特异性和对映立体选择性的新型麻黄碱脱氢酶的鉴定及研究

发布: 2021-12-24 572阅读 173评论

DOI: 10.1021/acscatal.9b00621

Tarek Shanati,Cameron Lockie,Lilian Beloti,Gideon Grogan,Marion B. Ansorge-Schumacher

2019年5月，德累斯顿工业大学（TUD）的Marion Ansorge-Schumacher教授及其团队从Arthrobacter sp. TS-15 (DSM 32400)鉴定分离出属于短链脱氢酶/还原酶（SDR）超家族的两个麻黄碱脱氢酶——PseDH和EDH。其中，PseDH和EDH催化1-苯基-1,2-丙二酮(1a)不对称还原分别生成(S)-1-羟基-1-苯基丙酮((S)-phenylacetylcarbinol,(S)-PAC)和(R)-PAC（如图1）。

图1  脱氢酶对1a的不对称氧化还原反应

作者探究并阐述了PseDH和EDH的功能和结构特征，首先是蛋白功能研究，包括两个脱氢酶的立体选择性、底物特异性以及酶动力学常数、蛋白质稳定性 。

底物特异性实验表明，PseDH和EDH能将多种芳基脂肪族羰基化合物(包括酮胺、酮酯和卤代酮)还原成相应的对映体醇；pH稳定性实验表明，PseDH和EDH分别在pH6.0−8.0和7.5−8.5范围内稳定性最好；热稳定性结果显示，在25℃时，PseDH比EDH更稳定，然而当温度升高至40℃时，EDH更稳定，其半衰期是25°C时的2倍。

基于上述结果，作者进一步解析了PseDH与辅酶NAD+复合物、底物1-苯基-1,2-丙二酮复合物的晶体结构（分辨率为1.83 Å），发现：PseDH具有SDR家族典型的关键催化残基——N113、S141、Y155 和K159。其中，K159与NAD+相互作用，降低了Y155的-OH基团的pKa值，从而引发了氢转移，即Y155在催化过程中作为“催化碱”存在，提供质子。被还原的α-羰基与Y155、S141存在相互作用，其中S141将羰基锚定在正确的还原方向上，起到固定底物的作用。同时，W152的侧链与底物1a的芳香环形成堆积作用。

图2  PseDH晶体结构

尽管决定立体选择性的活性位点在PseDH和EDH中尚未确定，但是研究者发现Y155和S141是PseDH中的保守氨基酸，在EDH中S143和W152分别被A和Y取代。将PseDH和EDH及其他30个不同来源的ADH氨基酸序列进行比对，结果表明：PseDH的氨基酸序列与EDH和RasADH的同源性最高（分别为32%和31.5%)，其中S143位常为小的疏水氨基酸如G、A或V，而W152位最常见的是H、M或L（图3）。因此，这两个位点的氨基酸残基在识别底物和定位还原反应中可能起关键作用。

图3  脱氢酶的多序列比对

新型脱氢酶PseDH和EDH是一种很有前途的绿色化学工具，它们优异的立体选择性和广泛的底物特异性为其在工业应用提供了光明的前景。此外，将提高PseDH和EDH稳定性作为新的研究起点，可进一步发挥其工业应用价值。

参考文献：

  1. Two Enantiocomplementary Ephedrine Dehydrogenases from Arthrobacter sp. TS-15 with Broad Substrate Specificity. DOI: 10.1021/acscatal.9b00621
  1. Critical residues for structure and catalysis in short-chain dehydrogenases/ reductases. DOI: 10.1074/jbc.M202160200
681赞
标签:



相关文章

发表回复

您的电子邮箱地址不会被公开。 必填项已用*标注

评论列表(173)

  19. “Back in the day, back before we turned into a modern society, a man would grab another man’s balls and swear an oath on them in his hand and state his words to be true. To be a friend” His father says. “Do you know what I mean, son?”

    回复

  33. I didn’t move and I didn’t want to move because the smell in there is so toxic. I can smell daddy’s unwashed ass in there. I was also so happy as daddy sat on the toilet to released his scent just for me to sniff. Just when The scent faded away I heard the daddy say, “Come here, boy.” I saw daddy sitting on my couch with nothing but a white brief on. The bulge was staring at me. I knew I was going to be having a wonderful night. “I think all of us were sprouting wood, today, dad.” He says. “Coach even noticed how hard we all were.”

    回复

  35. “I knew ya couldn’t keep your hands off it.” His dad says. “Men can’t do it, we are drawn to our cocks, like a moth to a flame, and usually that burning sensation that a man feels is the cum boiling up in our balls. You know that feelin’ doncha son?” His dad’s cock does not curve like that of his son, it angles like a fishing rod looking for fish under rushing river waters, from the deep furry patch between the father’s legs.

    回复

  37. I was surprised He wasn’t angry. He said,” boy you wait here, I am going to the bathroom.” He closed the door. He must sit on the toilet with pants down because I heard him losing the belt but didn’t hear him peeing. After a while, he called my name,” get in here ass sniffing slut.” I opened the door and crawled to the side of daddy. Daddy was still sitting on the toilet. . Just when I was confused why he sat that and did nothing. He stood up and pulled his pants up. I did take a peep of daddy’s huge uncut Middle Eastern dick. Half hard like 7inches already. While he was doing that, he grabbed my hair pushed right into the toilet, and closed the lid. “ smell my ass from there boy when it is still fresh.” He paused and continued, “ Don’t move.” Garrett picks up the pee-stained and cum-leaked jock from the tile floor.

    回复

  38. Regards, Wonderful information!
    milkyway casino online [url=https://usacasinomaster.com/#]free play online casino[/url] online cyprus casino

    回复

  39. “What…huh…what, dad?” He asks, puzzled by the question from his dad. As his cock is mere inches from his dad’s face. [url=https://arturzasada.pl/]porn sex[/url] Exactly a week after, daddy texted me with a different tone saying, “ get your ass ready tonight cause daddy is coming to your place tonight.” He was usually really sweet and nice, I was shot when I got the message. But automatically I said, “ yes daddy!” I guess I am a slutty whore for him right at the beginning as I knew my place where is always going to be inferior.

    回复

  68. Nicely put, Thanks a lot.
    online casino instant payout usa [url=https://usacasinomaster.com/#]no deposit bonus online casinos[/url] bovada online casino review

    回复
联系我们

联系我们

(44)07934433023

在线咨询: QQ交谈

邮箱: info@bioengx.org

关注微信
微信扫一扫关注我们

微信扫一扫关注我们

关注微博
返回顶部