Browsing by Author "Chang, Chin-Yuan"

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A study on the meta-data design for long-term digital multimedia preservation
(2008)
Chang, Feng-Cheng
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Chang, Chin-Yuan
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Hang, Hsueh-Ming
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電子工程學系及電子研究所
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Department of Electronics Engineering and Institute of Electronics
Due to the fast growth of multimedia content, the digital preservation emerges as an important technology that prolongs the life of digital objects. For audio-visual contents, we face the obsolescence problem in both supporting hardware/software and data representation formats. Migration strategy is a solution for achieving high-quality archival and easy access. In this paper, we investigate the concepts and structures in the Open Archival Information System (DAIS) and the MPEG-21 Digital Item Adaptation (DIA) specifications. By addressing the issues for a long-term audio-visual preservation system, we propose a combined meta-data scheme to record both the management and technical procedures and parameters. The detailed meta-data that keep the history of evolution versions can be used for subsequent migrations. We also implement a prototype system to realize our design. At the end of this study, we identify additional issues for future improvements.
Crystal Structure and Inhibition Studies of Transglutaminase from Streptomyces mobaraense
(2011-03-04)
Yang, Ming-Te
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Chang, Cheng-Hsiang
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Wang, Jiou Ming
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Wu, Tung Kung
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Wang, Yu-Kuo
The crystal structure of the microbial transglutaminase (MTGase) zymogen from Streptomyces mobaraense has been determined at 1.9-angstrom resolution using the molecular replacement method based on the crystal structure of the mature MTGase. The overall structure of this zymogen is similar to that of the mature form, consisting of a single disk-like domain with a deep active cleft at the edge of the molecule. A major portion of the prosequence (45 additional amino acid residues at the N terminus of the mature transglutaminase) folds into an L-shaped structure, consisting of an extended N-terminal segment linked with a one-turn short helix and a long alpha-helix. Two key residues in the short helix of the prosequence, Tyr-12 and Tyr-16, are located on top of the catalytic triad (Cys-110, Asp-301, and His-320) to block access of the substrate acyl donors and acceptors. Biochemical characterization of the mature MTGase, using N-alpha-benzyloxycarbonyl-L-glutaminylglycine as a substrate, revealed apparent K(m) and k(cat)/K(m) values of 52.66 mM and 40.42 mM(-1) min(-1), respectively. Inhibition studies using the partial prosequence SYAETYR and homologous sequence SQAETYR showed a noncompetitive inhibition mechanism with IC(50) values of 0.75 and 0.65 mM, respectively, but no cross-linking product formation. Nevertheless, the prosequence homologous oligopeptide SQAETQR, with Tyr-12 and Tyr-16 each replaced with Gln, exhibited inhibitory activity with the formation of the SQAETQR-monodansylcadaverine fluorophore cross-linking product (SQAETQR-C-DNS). MALDI-TOF tandem MS analysis of SQAETQR-C-DNS revealed molecular masses corresponding to those of (N)SQAETQ(C)-C-DNS and C-DNS-(N)QR(C) sequences, suggesting the incorporation of C-DNS onto the C-terminal Gln residue of the prosequence homologous oligopeptide. These results support the putative functional roles of both Tyr residues in substrate binding and inhibition.
Crystal Structure and Mutational Analysis of Aminoacylhistidine Dipeptidase from Vibrio alginolyticus Reveal a New Architecture of M20 Metallopeptidases
(2010-12-01)
Chang, Chin-Yuan
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Hsieh, Yin-Cheng
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Wang, Ting-Yi
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Chen, Yi-Chin
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Wang, Yu-Kuo
Aminoacylhistidine dipeptidases (PepD, EC 3.4.13.3) belong to the family of M20 metallopeptidases from the metallopeptidase H clan that catalyze a broad range of dipeptide and tripeptide substrates, including L-carnosine and L-homocarnosine. Homocarnosine has been suggested as a precursor for the neurotransmitter gamma-aminobutyric acid (GABA) and may mediate the antiseizure effects of GABAergic therapies. Here, we report the crystal structure of PepD from Vibrio alginolyticus and the results of mutational analysis of substrate-binding residues in the C-terminal as well as substrate specificity of the PepD catalytic domain-alone truncated protein PepD(CAT). The structure of PepD was found to exist as a homodimer, in which each monomer comprises a catalytic domain containing two zinc ions at the active site center for its hydrolytic function and a lid domain utilizing hydrogen bonds between helices to form the dimer interface. Although the PepD is structurally similar to PepV, which exists as a monomer, putative substrate-binding residues reside in different topological regions of the polypeptide chain. In addition, the lid domain of the PepD contains an "extra" domain not observed in related M20 family metallopeptidases with a dimeric structure. Mutational assays confirmed both the putative di-zinc allocations and the architecture of substrate recognition. In addition, the catalytic domain-alone truncated PepD(CAT) exhibited substrate specificity to L-homocarnosine compared with that of the wild-type PepD, indicating a potential value in applications of PepD(CAT) for GABAergic therapies or neuroprotection.
Crystal Structure and Mutational Analysis of Aminoacylhistidine Dipeptidase from Vibrio alginolyticus Reveal a New Architecture of M20 Metallopeptidases
(2010-12-01)
Chang, Chin-Yuan
;
Hsieh, Yin-Cheng
;
Wang, Ting-Yi
;
Chen, Yi-Chin
;
Wang, Yu-Kuo
Aminoacylhistidine dipeptidases (PepD, EC 3.4.13.3) belong to the family of M20 metallopeptidases from the metallopeptidase H clan that catalyze a broad range of dipeptide and tripeptide substrates, including L-carnosine and L-homocarnosine. Homocarnosine has been suggested as a precursor for the neurotransmitter gamma-aminobutyric acid (GABA) and may mediate the antiseizure effects of GABAergic therapies. Here, we report the crystal structure of PepD from Vibrio alginolyticus and the results of mutational analysis of substrate-binding residues in the C-terminal as well as substrate specificity of the PepD catalytic domain-alone truncated protein PepD(CAT). The structure of PepD was found to exist as a homodimer, in which each monomer comprises a catalytic domain containing two zinc ions at the active site center for its hydrolytic function and a lid domain utilizing hydrogen bonds between helices to form the dimer interface. Although the PepD is structurally similar to PepV, which exists as a monomer, putative substrate-binding residues reside in different topological regions of the polypeptide chain. In addition, the lid domain of the PepD contains an "extra" domain not observed in related M20 family metallopeptidases with a dimeric structure. Mutational assays confirmed both the putative di-zinc allocations and the architecture of substrate recognition. In addition, the catalytic domain-alone truncated PepD(CAT) exhibited substrate specificity to L-homocarnosine compared with that of the wild-type PepD, indicating a potential value in applications of PepD(CAT) for GABAergic therapies or neuroprotection.
Expression and characterization of the biofilm-related and carnosine-hydrolyzing aminoacylhistidine dipeptidase from Vibrio alginolyticus
(2008-10-01)
Wang, Ting-Yi
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Chen, Yi-Chin
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Kao, Liang-Wei
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Chang, Chin-Yuan
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Wang, Yu-Kuo
The biofilm-related and carnosine-hydrolyzing aminoacylhistidine dipeptidase (pepD) gene from Vibrio alginolyticus was cloned and sequenced. The recombinant PepD protein was produced and biochemically characterized and the putative active-site residues responsible for metal binding and catalysis were identified. The recombinant enzyme, which was identified as a homodimeric dipeptidase in solution, exhibited broad substrate specificity for Xaa-His and His-Xaa dipeptides, with the highest activity for the His-His dipeptide. Sequence and structural homologies suggest that the enzyme is a member of the metal-dependent metallopeptidase family. Indeed, the purified enzyme contains two zinc ions per monomer. Reconstitution of His.Tag-cleaved native apo-PepD with various metal ions indicated that enzymatic activity could be optimally restored when Zn(2+) was replaced with other divalent metal ions, including Mn(2+), Co(2+), Ni(2+), Cu(2+) and Cd(2+), and partially restored when Zn(2+) was replaced with Mg(2+). Structural homology modeling of PepD also revealed a 'catalytic domain' and a 'lid domain' similar to those of the Lactobacillus delbrueckii PepV protein. Mutational analysis of the putative active-site residues supported the involvement of His80, Asp119, Glu150, Asp173 and His461 in metal binding and Asp82 and Glu149 in catalysis. In addition, individual substitution of Glu149 and Glu150 with aspartic acid resulted in the partial retention of enzymatic activity, indicating a functional role for these residues on the catalysis and zinc ions, respectively. These effects may be necessary either for the activation of the catalytic water molecule or for the stabilization of the substrate-enzyme tetrahedral intermediate. Taken together, these results may facilitate the design of PepD inhibitors for application in antimicrobial treatment and antibody-directed enzyme prodrug therapy.
Genome mining and biosynthesis of kitacinnamycins as a STING activator
(2019-05-14)
Shi, Jing
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Liu, Cheng Li
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Zhang, Bo
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Guo, Wen Jie
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Zhu, Jiapeng
Cinnamoyl-containing nonribosomal peptides (CCNPs) are a small group of secondary metabolites with potent biological activities produced by actinobacteria. Two remarkable features in the biosynthesis of CCNPs include the nonribosomal peptide synthases (NRPSs) for assembly of the depsipeptide backbone and the type II polyketide synthases (PKSs) for N-terminal cinnamoyl moiety construction. Here, we present a genome mining approach targeting both NRPS and type II PKS for discovery of new CCNPs, which led to the identification of 51 putative CCNP gene clusters from public bacterial genome databases. After strain prioritization, a novel class of CCNP-type glycopeptides named kitacinnamycins, one of which showing potent activation ability towards the stimulator of interferon genes (STING) protein, was identified. Bioinformatic, genetic and biochemical analysis revealed the use of the NRPS assembly line to form the macrocyclic peptide backbone, followed by a P450 monooxygenase to generate terminal oxidized groups. A glycosyltransferase with relatively broad substrate specificity transfers sugars to the newly generated OH/COOH group. The protein crystallographic study further provided structural insights into this glycosylation. Our results not only demonstrated the feasibility of genome mining and strain prioritization for the discovery of new bioactive natural products but also disclosed the biosynthetic pathway for kitacinnamycins.
Multiple Pleomorphic Tetramers of Thermostable Direct Hemolysin from Grimontia hollisae in Exerting Hemolysis and Membrane Binding
(2019-07-08)
Wang, Yu-Kuo
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Huang, Sheng-Cih
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Chang, Chin-Yuan
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Huang, Wan-Ting
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Liao, Man-Jun
Oligomerization of protein into specific quaternary structures plays important biological functions, including regulation of gene expression, enzymes activity, and cell-cell interactions. Here, we report the determination of two crystal structures of the Grimontia hollisae (formally described as Vibrio hollisae) thermostable direct hemolysin (Gh-TDH), a pore-forming toxin. The toxin crystalized in the same space group of P2(1)2(1)2, but with two different crystal packing patterns, each revealing three consistent tetrameric oligomerization forms called Oligomer-I, -II, and -III. A central pore with comparable depth of similar to 50 angstrom but differing in shape and size was observed in all determined toxin tetrameric oligomers. A common motif of a toxin dimer was found in all determined structures, suggesting a plausible minimum functional unit within the tetrameric structure in cell membrane binding and possible hemolytic activity. Our results show that bacterial toxins may form a single or highly symmetric oligomerization state when exerting their biological functions. The dynamic nature of multiple symmetric oligomers formed upon release of the toxin may open a niche for bacteria survival in harsh living environments.
Pregnenolonyl-alpha-glucoside exhibits marked anti-cancer and CYP17A1 enzymatic inhibitory activities
(2020-02-07)
Chou, Feng-Pai
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Hsu, Wen-Chen
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Huang, Sheng-Cih
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Chang, Chin-Yuan
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Chiou, Ya-Sheng
We report here that pregnenolonyl-alpha-glucoside (2), a steryl glycoside synthesized directly from pregnenolone and glucose via a consecutive multienzyme-catalyzed process, exhibits marked dose-dependent cytotoxic activity against HT29, AGS, and ES-2 cells with IC50 values of 23.5 to 50.9 mu M. An in vitro CYP17A1 binding pattern assay and protein-ligand docking model support that 2, like abiraterone, binds in the active site heme iron pocket of CYP17A1.
Purification, crystallization and preliminary X-ray analysis of an aminoacylhistidine dipeptidase (PepD) from Vibrio alginolyticus
(2009-03-01)
Chang, Chin-Yuan
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Hsieh, Yin-Cheng
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Wang, Ting-Yi
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Chen, Chun-Jung
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Wu, Tung-Kung
The aminoacylhistidine dipeptidase (PepD) protein encoded by Vibrio alginolyticus pepD was successfully overexpressed and characterized and the putative active-site residues responsible for metal binding and catalysis were identified. The purified enzyme contained two zinc ions per monomer. The recombinant dipeptidase enzyme, which was identified as a homodimer in solution, exhibited broad substrate specificity for Xaa-His dipeptides, with highest activity towards the His-His dipeptide. The purified protein was crystallized using the hanging-drop vapour-diffusion method. Preliminary crystallographic analysis showed that the crystal belonged to space group P6(1) or P6(5), with unit-cell parameters a = b = 80.42, c = 303.11 angstrom. The crystal contained two molecules per asymmetric unit and the predicted solvent content was 53.4%.
Terpene synthases in disguise: enzymology, structure, and opportunities of non-canonical terpene synthases
(2020-03-01)
Rudolf, Jeffrey D.
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Chang, Chin-Yuan
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生物科技學系
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Department of Biological Science and Technology
Covering: up to July 2019 Terpene synthases (TSs) are responsible for generating much of the structural diversity found in the superfamily of terpenoid natural products. These elegant enzymes mediate complex carbocation-based cyclization and rearrangement cascades with a variety of electron-rich linear and cyclic substrates. For decades, two main classes of TSs, divided by how they generate the reaction-triggering initial carbocation, have dominated the field of terpene enzymology. Recently, several novel and unconventional TSs that perform TS-like reactions but do not resemble canonical TSs in sequence or structure have been discovered. In this review, we identify 12 families of non-canonical TSs and examine their sequences, structures, functions, and proposed mechanisms. Nature provides a wide diversity of enzymes, including prenyltransferases, methyltransferases, P450s, and NAD(+)-dependent dehydrogenases, as well as completely new enzymes, that utilize distinctive reaction mechanisms for TS chemistry. These unique non-canonical TSs provide immense opportunities to understand how nature evolved different tools for terpene biosynthesis by structural and mechanistic characterization while affording new probes for the discovery of novel terpenoid natural products and gene clusters via genome mining. With every new discovery, the dualistic paradigm of TSs is contradicted and the field of terpene chemistry and enzymology continues to expand.