2IUX

Structure of Testis Ace Glycosylation Mutants and Evidence for Conserved Domain Movement.

(2006) Biochemistry 45: 12654

• PubMed: 17042482 Search on PubMedSearch on PubMed Central
• DOI: https://doi.org/10.1021/bi061146z
• Primary Citation of Related Structures:  
  2IUL, 2IUX


• PubMed Abstract: 
  

  Human angiotensin-converting enzyme is an important drug target for which little structural information has been available until recent years. The slow progress in obtaining a crystal structure was due to the problem of surface glycosylation, a difficulty that has thus far been overcome by the use of a glucosidase-1 inhibitor in the tissue culture medium. However, the prohibitive cost of these inhibitors and incomplete glucosidase inhibition makes alternative routes to minimizing the N-glycan heterogeneity desirable. Here, glycosylation in the testis isoform (tACE) has been reduced by Asn-Gln point mutations at N-glycosylation sites, and the crystal structures of mutants having two and four intact sites have been solved to 2.0 A and 2.8 A, respectively. Both mutants show close structural identity with the wild-type. A hinge mechanism is proposed for substrate entry into the active cleft, based on homology to human ACE2 at the levels of sequence and flexibility. This is supported by normal-mode analysis that reveals intrinsic flexibility about the active site of tACE. Subdomain II, containing bound chloride and zinc ions, is found to have greater stability than subdomain I in the structures of three ACE homologues. Crystallizable glycosylation mutants open up new possibilities for cocrystallization studies to aid the design of novel ACE inhibitors.

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Organizational Affiliation: 

Division of Medical Biochemistry, Institute of Infectious Disease and Molecular Medicine, University of Cape Town, South Africa.