Structure of testis ACE glycosylation mutants and evidence for conserved domain movement
| dc.contributor.author | Watermeyer, Jean M | |
| dc.contributor.author | Sewell, Trevor B | |
| dc.contributor.author | Schwager, Sylva L | |
| dc.contributor.author | Natesh, Ramanathan | |
| dc.contributor.author | Corradi, Hazel R | |
| dc.contributor.author | Acharya, Ravi K | |
| dc.contributor.author | Sturrock, Edward D | |
| dc.date.accessioned | 2016-07-28T14:37:39Z | |
| dc.date.available | 2016-07-28T14:37:39Z | |
| dc.date.issued | 2006 | |
| dc.date.updated | 2016-07-28T14:31:47Z | |
| dc.description.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 Å and 2.8 Å, 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. | en_ZA |
| dc.identifier | http://dx.doi.org/10.1021/bi061146z | |
| dc.identifier.apacitation | Watermeyer, J. M., Sewell, T. B., Schwager, S. L., Natesh, R., Corradi, H. R., Acharya, R. K., & Sturrock, E. D. (2006). Structure of testis ACE glycosylation mutants and evidence for conserved domain movement. <i>Biochemistry</i>, http://hdl.handle.net/11427/21004 | en_ZA |
| dc.identifier.chicagocitation | Watermeyer, Jean M, Trevor B Sewell, Sylva L Schwager, Ramanathan Natesh, Hazel R Corradi, Ravi K Acharya, and Edward D Sturrock "Structure of testis ACE glycosylation mutants and evidence for conserved domain movement." <i>Biochemistry</i> (2006) http://hdl.handle.net/11427/21004 | en_ZA |
| dc.identifier.citation | Watermeyer, J. M., Sewell, B. T., Schwager, S. L., Natesh, R., Corradi, H. R., Acharya, K. R., & Sturrock, E. D. (2006). Structure of testis ACE glycosylation mutants and evidence for conserved domain movement. Biochemistry, 45(42), 12654-12663. | en_ZA |
| dc.identifier.issn | 0006-2960 | en_ZA |
| dc.identifier.ris | TY - Journal Article AU - Watermeyer, Jean M AU - Sewell, Trevor B AU - Schwager, Sylva L AU - Natesh, Ramanathan AU - Corradi, Hazel R AU - Acharya, Ravi K AU - Sturrock, Edward D AB - 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 Å and 2.8 Å, 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. DA - 2006 DB - OpenUCT DP - University of Cape Town J1 - Biochemistry LK - https://open.uct.ac.za PB - University of Cape Town PY - 2006 SM - 0006-2960 T1 - Structure of testis ACE glycosylation mutants and evidence for conserved domain movement TI - Structure of testis ACE glycosylation mutants and evidence for conserved domain movement UR - http://hdl.handle.net/11427/21004 ER - | en_ZA |
| dc.identifier.uri | http://hdl.handle.net/11427/21004 | |
| dc.identifier.vancouvercitation | Watermeyer JM, Sewell TB, Schwager SL, Natesh R, Corradi HR, Acharya RK, et al. Structure of testis ACE glycosylation mutants and evidence for conserved domain movement. Biochemistry. 2006; http://hdl.handle.net/11427/21004. | en_ZA |
| dc.language | eng | en_ZA |
| dc.publisher | American Chemical Society | en_ZA |
| dc.publisher.institution | University of Cape Town | |
| dc.source | Biochemistry | en_ZA |
| dc.source.uri | http://pubs.acs.org/journal/bichaw | |
| dc.title | Structure of testis ACE glycosylation mutants and evidence for conserved domain movement | en_ZA |
| dc.type | Journal Article | en_ZA |
| uct.type.filetype | Text | |
| uct.type.filetype | Image | |
| uct.type.publication | Research | en_ZA |
| uct.type.resource | Article | en_ZA |