The mechanism of thia-Michael addition catalyzed by LanC enzymes

Author:

Ongpipattanakul Chayanid1ORCID,Liu Shi2ORCID,Luo Youran2ORCID,Nair Satish K.123ORCID,van der Donk Wilfred A.12345ORCID

Affiliation:

1. Department of Biochemistry, University of Illinois at Urbana-Champaign, Urbana, IL 61801

2. Department of Chemistry, University of Illinois at Urbana-Champaign, Urbana, IL 61801

3. Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801

4. HHMI, University of Illinois at Urbana-Champaign, Urbana, IL 61801

5. Center for Biophysics and Computational Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801

Abstract

In both eukarya and bacteria, the addition of Cys to dehydroalanine (Dha) and dehydrobutyrine (Dhb) occurs in various biological processes. In bacteria, intramolecular thia-Michael addition catalyzed by lanthipeptide cyclases (LanC) proteins or protein domains gives rise to a class of natural products called lanthipeptides. In eukarya, dehydroamino acids in signaling proteins are introduced by effector proteins produced by pathogens like Salmonella to dysregulate host defense mechanisms. A eukaryotic LanC-like (LanCL) enzyme catalyzes the addition of Cys in glutathione to Dha/Dhb to protect the cellular proteome from unwanted chemical and biological activity. To date, the mechanism of the enzyme-catalyzed thia-Michael addition has remained elusive. We report here the crystal structures of the human LanCL1 enzyme complexed with different ligands, including the product of thia-Michael addition of glutathione to a Dhb-containing peptide that represents the activation loop of Erk. The structures show that a zinc ion activates the Cys thiolate for nucleophilic attack and that a conserved His is poised to protonate the enolate intermediate to achieve a net anti- addition. A second His hydrogen bonds to the carbonyl oxygen of the former Dhb and may stabilize the negative charge that builds up on this oxygen atom in the enolate intermediate. Surprisingly, the latter His is not conserved in orthologous enzymes that catalyze thia-Michael addition to Dha/Dhb. Eukaryotic LanCLs contain a His, whereas bacterial stand-alone LanCs have a Tyr residue, and LanM enzymes that have LanC-like domains have a Lys, Asn, or His residue. Mutational and binding studies support the importance of these residues for catalysis.

Funder

HHS | NIH | National Institute of General Medical Sciences

Publisher

Proceedings of the National Academy of Sciences

Subject

Multidisciplinary

同舟云学术

1.学者识别学者识别

2.学术分析学术分析

3.人才评估人才评估

"同舟云学术"是以全球学者为主线,采集、加工和组织学术论文而形成的新型学术文献查询和分析系统,可以对全球学者进行文献检索和人才价值评估。用户可以通过关注某些学科领域的顶尖人物而持续追踪该领域的学科进展和研究前沿。经过近期的数据扩容,当前同舟云学术共收录了国内外主流学术期刊6万余种,收集的期刊论文及会议论文总量共计约1.5亿篇,并以每天添加12000余篇中外论文的速度递增。我们也可以为用户提供个性化、定制化的学者数据。欢迎来电咨询!咨询电话:010-8811{复制后删除}0370

www.globalauthorid.com

TOP

Copyright © 2019-2024 北京同舟云网络信息技术有限公司
京公网安备11010802033243号  京ICP备18003416号-3