Abstract

Mutagenic studies that implicate specific amino acids in reaction chemistry or in substrate binding are fully interpretable only when the structural integrity of the variant protein has been validated. This validation is facilitated by identification of spin-labeled substrate analogs. Measurement of spin-probe affinity and binding stoichiometry, using a C266S and C266A mutants and comparison of these parameters with those measured for wild-type protein affords an elegant, rigorous, yet efficient solution-state method for validating the structural integrity of engineered proteins. The paramagnetic cation Mn++ is being productively used to validate the integrity of engineered variants of human HMG-CoA lyase, which catalyzed a key step in leucine catabolism. Studies were conducted using both X- and Q-band EPR spectroscopy.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR001008-21
Application #
5222125
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
21
Fiscal Year
1996
Total Cost
Indirect Cost

  Publications

Mao, Li; Liu, Yu-Xiang; Huang, Chun-Hua et al. (2015) Intrinsic Chemiluminescence Generation during Advanced Oxidation of Persistent Halogenated Aromatic Carcinogens. Environ Sci Technol 49:7940-7
Shan, Guo-Qiang; Yu, Ao; Zhao, Chuan-Fang et al. (2015) A combined experimental and computational investigation on the unusual molecular mechanism of the Lossen rearrangement reaction activated by carcinogenic halogenated quinones. J Org Chem 80:180-9
Li, Yan; Huang, Chun-Hua; Liu, Yu-Xiang et al. (2014) Detoxifying polyhalogenated catechols through a copper-chelating agent by forming stable and redox-inactive hydrogen-bonded complexes with an unusual perpendicular structure. Chemistry 20:13028-33
Shao, Jie; Huang, Chun-Hua; Kalyanaraman, Balaraman et al. (2013) Potent methyl oxidation of 5-methyl-2'-deoxycytidine by halogenated quinoid carcinogens and hydrogen peroxide via a metal-independent mechanism. Free Radic Biol Med 60:177-82
Sheng, Zhi-Guo; Li, Yan; Fan, Rui-Mei et al. (2013) Lethal synergism between organic and inorganic wood preservatives via formation of an unusual lipophilic ternary complex. Toxicol Appl Pharmacol 266:335-44
Qin, Hao; Huang, Chun-Hua; Mao, Li et al. (2013) Molecular mechanism of metal-independent decomposition of lipid hydroperoxide 13-HPODE by halogenated quinoid carcinogens. Free Radic Biol Med 63:459-66
Huang, Chun-Hua; Shan, Guo-Qiang; Mao, Li et al. (2013) The first purification and unequivocal characterization of the radical form of the carbon-centered quinone ketoxy radical adduct. Chem Commun (Camb) 49:6436-8
Sheng, Zhi-Guo; Huang, Wei; Liu, Yu-Xiang et al. (2013) Ofloxacin induces apoptosis via ?1 integrin-EGFR-Rac1-Nox2 pathway in microencapsulated chondrocytes. Toxicol Appl Pharmacol 267:74-87
Sheng, Zhi-Guo; Huang, Wei; Liu, Yu-Xiang et al. (2013) Bisphenol A at a low concentration boosts mouse spermatogonial cell proliferation by inducing the G protein-coupled receptor 30 expression. Toxicol Appl Pharmacol 267:88-94
Liddle, Brendan J; Wanniarachchi, Sarath; Hewage, Jeewantha S et al. (2012) Electronic communication across diamagnetic metal bridges: a homoleptic gallium(III) complex of a redox-active diarylamido-based ligand and its oxidized derivatives. Inorg Chem 51:12720-8

Showing the most recent 10 out of 368 publications