Antiquitin (ATQ) belongs to the superfamily of aldehyde dehydrogenase (ALDH). It is an evolutionarily conserved protein as shown from its high amino acid sequence identity between human and its plant counterparts. Therefore, ATQ is believed to play an important physiological role. Until now, however, studies on ATQ are limited and its cellular function is uncertain. Recently, we have first demonstrated the aldehyde oxidizing ability of ATQ purified from the liver of black seabream (Acanthopagrus schlegeli). To further investigate this protein, different attempts have been made. / Recombinant ATQ has been successfully expressed in E. coli. Kinetics studies showed that it possessed similar characteristics with its native enzyme. The recombinant protein was produced in large amount for protein crystallization. Crystal of ATQ was obtained and its X-ray structure was solved to 2.8 A in complex with NAD+. Tetrameric ATQ was a dimer of dimer. Three domains can be found in the subunit structure of ATQ, the NAD+-binding domain, catalytic domain and oligomerization domain. In each of the NAD+-binding domain, one molecule of NAD + could be found. The overall structure of ATQ was similar to other tetrameric ALDHs, but the coenzyme binding was in a single "hydride transfer" conformation and the density was well-defined which was contrast to most ALDH structures. Structural study of the substrate-binding pocket explained the failure of ATQ in oxidizing several aldehydes which is specific to certain members of ALDH. / The ATQ full-length cDNA of black seabream was obtained. It consisted of 2309 by with a 153 nucleotide long 5' UTR, and a 209 nucleotide long 3' UTR. An ORF of 1533 by which encoded a protein with 511 amino acids was found. This putative protein showed the highest of 87% sequence identity with zebrafish ATQ, and ∼60% with plant ATQs. Tissue distribution was studied by RT-PCR. A high level of mRNA expression was observed in liver and kidney. Subcellular localization study using green fluorescent protein (GFP) fusion protein showed that ATQ was expressed in cytoplasm. However, another in-frame initiation methionine (M1) was found 31 residues before this generally accepted methionine (M2). Both iPSORT analysis and experimental studies using GFP fusion protein indicated that the 31 amino acid peptide contained a mitochondrial-targeting signal. / Tang Wai Kwan. / "July 2005." / Adviser: Fong Wing Ping. / Source: Dissertation Abstracts International, Volume: 67-07, Section: B, page: 3603. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (p. 130-145). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract in English and Chinese. / School code: 1307.
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_343672 |
| Date | January 2005 |
| Contributors | Tang, Wai Kwan., Chinese University of Hong Kong Graduate School. Division of Biochemistry. |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, theses |
| Format | electronic resource, microform, microfiche, 1 online resource (viii, 149 p. : ill.) |
| Rights | Use of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/) |
Page generated in 0.0023 seconds