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This content will become publicly available on December 22, 2018

Title: Structural Evidence of a Major Conformational Change Triggered by Substrate Binding in DapE Enzymes: Impact on the Catalytic Mechanism

In this paper, the X-ray crystal structure of the dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase from Haemophilus influenzae (HiDapE) bound by the products of hydrolysis, succinic acid and l,l-DAP, was determined at 1.95 Å. Surprisingly, the structure bound to the products revealed that HiDapE undergoes a significant conformational change in which the catalytic domain rotates ~50° and shifts ~10.1 Å (as measured at the position of the Zn atoms) relative to the dimerization domain. This heretofore unobserved closed conformation revealed significant movements within the catalytic domain compared to that of wild-type HiDapE, which results in effectively closing off access to the dinuclear Zn(II) active site with the succinate carboxylate moiety bridging the dinculear Zn(II) cluster in a μ-1,3 fashion forming a bis(μ-carboxylato)dizinc(II) core with a Zn–Zn distance of 3.8 Å. Surprisingly, His194.B, which is located on the dimerization domain of the opposing chain ~10.1 Å from the dinuclear Zn(II) active site, forms a hydrogen bond (2.9 Å) with the oxygen atom of succinic acid bound to Zn2, forming an oxyanion hole. As the closed structure forms upon substrate binding, the movement of His194.B by more than ~10 Å is critical, based on site-directed mutagenesis data, for activation of the scissile carbonyl carbonmore » of the substrate for nucleophilic attack by a hydroxide nucleophile. Employing the HiDapE product-bound structure as the starting point, a reverse engineering approach called product-based transition-state modeling provided structural models for each major catalytic step. Finally, these data provide insight into the catalytic reaction mechanism and also the future design of new, potent inhibitors of DapE enzymes.« less
Authors:
 [1] ;  [2] ;  [2] ;  [2] ;  [3] ;  [1] ;  [1] ;  [1] ;  [2] ; ORCiD logo [3]
  1. Argonne National Lab. (ANL), Argonne, IL (United States). Midwest Center for Structural Genomics. Structural Biology Center. Biosciences Division
  2. Loyola Univ. Chicago, IL (United States). Dept. of Chemistry and Biochemistry
  3. Marquette Univ., Milwaukee, WI (United States). Dept. of Chemistry
Publication Date:
Grant/Contract Number:
AC02-06CH11357; HHSN272200700058C; HHSN272201200026C; CHE-1412443
Type:
Accepted Manuscript
Journal Name:
Biochemistry
Additional Journal Information:
Journal Volume: 57; Journal Issue: 5; Journal ID: ISSN 0006-2960
Publisher:
American Chemical Society (ACS)
Research Org:
Argonne National Lab. (ANL), Argonne, IL (United States); Loyola Univ. Chicago, IL (United States); Marquette Univ., Milwaukee, WI (United States)
Sponsoring Org:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23); National Inst. of Health (NIH) (United States); National Science Foundation (NSF); Todd Wehr Foundation (United States)
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY
OSTI Identifier:
1427504