Browsing by Author "Chang, Cheng-Hsiang"

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A Compact 4.4 Tesla Cryogenic Wavelength Shifter for the Taiwan Photon Source
(2017)
Wang, Chao-Chin
;
Chang, Cheng-Hsiang
;
Hwang, Ching-Shiang
;
Lee, Chih-Hao
;
電子物理學系
The feasibility of a compact 4.4 T in-vacuum cryogenic Pr2Fe14B-based wavelength shifter was studied at the Taiwan Photon Source. The overall dimensions of the magnet array are height 205 mm, length 240 mm, and width 160 mm; the magnet array operates at magnet gap 5 mm. The roll-off of the central field on the transverse axis was designed to be less than 0.1% between +/- 20 mm. The magnitudes of the integral multipole field error and the transverse tune shift as a function of transverse position were optimized to be less than 200 G cm and 4 x 10(-3) between +/- 15 mm. A reverse field occurred so that the Pr-Fe-B bulk became partially demagnetized near 295 K, but it was improved upon chamfering the iron poles near the regions of that reverse field to avoid demagnetization of the Pr-Fe-B bulk.
A Compact 4.4 Tesla Cryogenic Wavelength Shifter for the Taiwan Photon Source
(2017)
Wang, Chao-Chin
;
Chang, Cheng-Hsiang
;
Hwang, Ching-Shiang
;
Lee, Chih-Hao
;
電子物理學系
The feasibility of a compact 4.4 T in-vacuum cryogenic Pr2Fe14B-based wavelength shifter was studied at the Taiwan Photon Source. The overall dimensions of the magnet array are height 205 mm, length 240 mm, and width 160 mm; the magnet array operates at magnet gap 5 mm. The roll-off of the central field on the transverse axis was designed to be less than 0.1% between +/- 20 mm. The magnitudes of the integral multipole field error and the transverse tune shift as a function of transverse position were optimized to be less than 200 G cm and 4 x 10(-3) between +/- 15 mm. A reverse field occurred so that the Pr-Fe-B bulk became partially demagnetized near 295 K, but it was improved upon chamfering the iron poles near the regions of that reverse field to avoid demagnetization of the Pr-Fe-B bulk.
Active and passive compensation of APPLE II-introduced multipole errors through beam-based measurement
(2016-08-01)
Chung, Ting-Yi
;
Huang, Szu-Jung
;
Fu, Huang-Wen
;
Chang, Ho-Ping
;
Chang, Cheng-Hsiang
The effect of an APPLE II-type elliptically polarized undulator (EPU) on the beam dynamics were investigated using active and passive methods. To reduce the tune shift and improve the injection efficiency, dynamic multipole errors were compensated using L-shaped iron shims, which resulted in stable top-up operation for a minimum gap. The skew quadrupole error was compensated using a multipole corrector, which was located downstream of the EPU for minimizing betatron coupling, and it ensured the enhancement of the synchrotron radiation brightness. The investigation methods, a numerical simulation algorithm, a multipole error correction method, and the beam-based measurement results are discussed. (C) 2016 Elsevier B.V. All rights reserved.
Alteration of the Substrate's Prefolded Conformation and Cyclization Stereochemistry of Oxidosqualene-Lanosterol Cyclase of Saccharomyces cerevisiae by Substitution at Phenylalanine 699
(2010-02-05)
Wu, Tung-Kung
;
Chang, Cheng-Hsiang
;
Wen, Hao-Yu
;
Liu, Yuan-Ting
;
Li, Wen-Hsuan
The Saccharomyces cerevisiae ERG7(Phe699) mutants produced one chair-chair-chair (C-C-C) and two chair-boat-chair (C-B-C) truncated tricyclic compounds, one tetracyclic 17 alpha-exocyclic unrearranged intermediate, and two 17 beta-exocyclic truncated rearranged intermediates. These results provided direct evidence for the importance of the residue in affecting mechanistic transitions between C-B-C and C-C-C substrate conformation and between the 17 alpha- and 17 beta-exocyclic side chain stereochemistry as well as in stabilizing the 6 -6-5 tricyclic and the protosteryl C-17 cations.
Crystal Structure and Inhibition Studies of Transglutaminase from Streptomyces mobaraense
(2011-03-04)
Yang, Ming-Te
;
Chang, Cheng-Hsiang
;
Wang, Jiou Ming
;
Wu, Tung Kung
;
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
;
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.
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.
Differential Stereocontrolled Formation of Tricyclic Triterpenes by Mutation of Tyrosine 99 of the Oxidosqualene-Lanosterol Cyclase from Saccharomyces cerevisiae
(2009-11-01)
Wu, Tung-Kung
;
Li, Wen-Hsuan
;
Chang, Cheng-Hsiang
;
Wen, Hao-Yu
;
Liu, Yuan-Ting
The function of the Tyr99 residue from Saccharomyces cerevisiae oxidosqualene-lanosterol cyclase (ERG7) was analyzed by constructing deletion and site-saturated mutants. Two truncated intermediates, (13 alpha H)-isomalabarica-14Z,17E,21-trien-3 beta-ol and (13 alpha H)-isomalabarica-14E,17E,21-trien-3 beta-ol, were isolated from the ERG7(Y99X) mutants. These results suggest that the functional role of ERG7(Y99) is to affect both chair-boat 6-6-5 tricyclic Markovnikov C-14 cation stabilization and the stereochemistry of the protons at the C-15 position for subsequent deprotonation. ((C) Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2009)
Importance of Saccharomyces cerevisiae Oxidosqualene-Lanosterol Cyclase Tyrosine 707 Residue for Chair-Boat Bicyclic Ring Formation and Deprotonation Reactions
(2008-11-06)
Wu, Tung-Kung
;
Wang, Tsai-Ting
;
Chang, Cheng-Hsiang
;
Liu, Yuan-Ting
;
Shie, Wen-Shiang
A contact mapping strategy was applied to identity putative amino acid residues that influence the oxidosqualene-lanosterol B-ring cyclization reaction. A bicyclic intermediate with two altered deprotonation products, in conjunction with lanosterol, were isolated from the ERG7(Y707X) mutants, indicating that the Tyr707 residue may play a functional role in stabilizing the chair-boat bicyclic C-8 cation and the lanosteryl C-8/C-9 cation intermediates.
Mutation of isoleucine 705 of the oxidosqualene-lanosterol cyclase from Saccharomyces cerevisiae affects lanosterol's C/D-ring cyclization and 17 alpha/beta-exocyclic side chain stereochemistry
(2011)
Wu, Tung-Kung
;
Chang, Yi-Chun
;
Liu, Yuan-Ting
;
Chang, Cheng-Hsiang
;
Wen, Hao-Yu
Site-saturated substitution in Saccharomyces cerevisiae oxidosqualene-lanosterol cyclase at Ile705 position produced three chair-boat-chair (C-B-C) truncated tricyclic compounds, two 17 alpha-exocyclic protosteryl intermediates, two protosteryl C-17 truncated rearranged intermediates and the normal biosynthetic product, lanosterol. These results indicated the importance of the Ile705 residue in affecting lanosterol's C/D ring stabilization including 6-6-5 tricyclic and protosteryl C-17 cations and 17 alpha/beta-exocyclic side chain stereochemistry.
ORGN 611-Molecular cloning and inhibition studies of oxidosqualene-lanosterol cyclase from bovine liver using a novel Ro48-8071 based fluorescence probe
(2006-09-10)
Chang, Cheng-Hsiang
;
Wu, Tung-Kung
;
生物科技學系
;
Department of Biological Science and Technology
Phenylalanine 445 within oxidosqualene-lanosterol cyclase from Saccharomyces cerevisiae influences C-ring cyclization and deprotonation reactions
(2006-10-12)
Wu, Tung-Kung
;
Liu, Yuan-Ting
;
Chiu, Feng-Hsuan
;
Chang, Cheng-Hsiang
;
生物科技學系
We describe the Saccharomyces cerevisiae oxidosqualene - lanosterol cyclase Phe445 site-saturated mutants that generate truncated tricyclic and altered deprotonation product profiles. Among these mutants, only polar side-chain group substitutions genetically complemented yeast viability and produced spatially related product diversity, supporting the Johnson model that cation - pi interactions between a carbocationic intermediate and an enzyme can be replaced by an electrostatic or polar side chain to stabilize the cationic intermediate, but with product differentiation.
Photoactivation Studies of Zinc Porphyrin-Myoglobin System and Its Application for Light-Chemical Energy Conversion
(2011)
Chang, Chin-Hao
;
Hu, Yi-Ting
;
Lo, Chen-Fu
;
Luo, Liyang
;
Lin, Hung-Ming
An artificial zinc porphyrin-myoglobin- based photo-chemical energy conversion system, consisting of ZnPP-Mb or ZnPE1-Mb as a photosensitizer, NADP(+) as an electron acceptor, and triethanolamine as an electron donor, has been constructed to mimic photosystem I. The photoirradiated product is able to reduce a single-electron acceptor protein cytochrome c, but cannot catalyze the two-electron reduction of acetaldehyde by alcohol dehydrogenase, thus demonstrating a single electron transfer mechanism. Furthermore, the artificial system can bifunctionally promote oxidoredox reactions, depending on the presence or absence of a sacrificial electron donor, thus suggesting its potential application in electrochemical regeneration steps involved in chemical transformation and/or energy conversion.
Protein engineering of oxidosqualene-lanosterol cyclase into triterpene monocyclase
(2013)
Chang, Cheng-Hsiang
;
Wen, Hao-Yu
;
Shie, Wen-Shiang
;
Lu, Ching-Ting
;
Li, Meng-Erh
A computational modeling/protein engineering approach was applied to probe H234, C457, T509, Y510, and W587 within Saccharomyces cerevisiae oxidosqualene-lanosterol cyclase (ERG7), which spatially affects the C-10 cation of lanosterol formation. Substitution of Trp587 to aromatic residues supported the "aromatic hypothesis" that the pi-electron-rich pocket is important for the stabilization of electron-deficient cationic intermediates. The Cys457 to Gly and Thr509 to Gly mutations disrupted the pre-existing H-bond to the protonating Asp456 and the intrinsic His234 : Tyr510 H-bond network, respectively, and generated achilleol A as the major product. An H234W/Y510W double mutation altered the ERG7 function to achilleol A synthase activity and generated achilleol A as the sole product. These results support the concept that a few-ring triterpene synthase can be derived from polycyclic cyclases by reverse evolution, and exemplify the power of computational modeling coupled with protein engineering both to study the enzyme's structure-function-mechanism relationships and to evolve new enzymatic activity.
Protein Engineering of Saccharomyces cerevisiae Oxidosqualene-Lanosterol Cyclase into Parkeol Synthase
(2012-10-19)
Liu, Yuan-Ting
;
Hu, Tain-Chang
;
Chang, Cheng-Hsiang
;
Shie, Wen-Shiang
;
Wu, Tung-Kung
A Saccharomyces cerevisiae oxidosqualene-lanosterol cyclase mutant, ERG7(T384Y/Q450H/V4541), produced parkeol but not lanosterol as the sole end product. Parkeol undergoes downstream metabolism to generate compounds 9 and 10. In vitro incubation of parkeol produced a product profile similar to that of the in vivo experiment. In summary, parkeol undergoes a metabolic pathway similar to that of cycloartenol in yeast but distinct from that of lanosterol in yeast, suggesting that two different metabolic pathways of postoxidosqualene cyclization may exist in S. cerevisiae.
Protein plasticity: A single amino acid substitution in the Saccharomyces cerevisiae oxidosqualene-lanosterol cyclase generates protosta-13(17),24-dien-3 beta-ol, a rearrangement product
(2008-06-19)
Wu, Tung-Kung
;
Wen, Hao-Yu
;
Chang, Cheng-Hsiang
;
Liu, Yuan-Ting
;
應用化學系
To provide insights into the structure-function relationships of oxidosqualene-lanosterol cyclase (ERG7) from Saccharomyces cerevisiae, the Phe699 was exchanged against hydrophilic polar uncharged residues Ser, Thr, Cys, Gin, and Tyr to characterize its product profile and functional role in ERG7 activity. Among the substitutions, only the ERG7(F699T) mutant produced novel protosta-13(17),24-dien-3 beta-ol as the sole truncated rearrangement product. The results suggest that Phe699Thr mutation is likely to affect the C-17 cation stabilization during the rearrangement process.
Purification and Properties of a Novel Phenolic Antioxidant from Radix astragali Fermented by Aspergillus oryzae M29
(2011-06-22)
Sheih, I-Chuan
;
Fang, Tony J.
;
Wu, Tung-Kung
;
Chang, Cheng-Hsiang
;
Chen, Ru-Yin
The Chinese herb Radix astragalus (RA) has been widely used as a dietary supplement in Asia, and there are numerous reports on its bioactivities. However, there are no reports to date regarding the use of Aspergillus spp. in the culture medium of the RA plant for the production of phenolic antioxidants. In this study, utilizing the fungus Aspergillus to ferment the native RA has successfully resulted in a significant increase in the phenolic contents of RA, and the fermented RA also revealed much better antioxidant activity toward 2,2'-azinobis (3-ethylbenzothiazoline-6-sulfonic acid (ABTS) radicals, hydroxyl radical, superoxide radical and peroxyl radical than those of unfermented RA. Among these phenolics, a potent novel antioxidant was isolated and identified as 3,4-di(4'-hydroxyphenyl) isobutyric acid with a molecular weight of 272, by ESI-MS (electrospray ionization mass), (1)H NMR (nuclear magnetic resonance), (13)C NMR, DEPT (distortionless enhancement by polarization transfer)-NMR, HMQC (heteronuclear multiple quantum coherence), and HMBC (heteronuclear multiple bond correlation) spectra. These data demonstrated that the solid-state bioprocessing strategy could be an innovative approach to enhance the antioxidant activity of RA.
Saccharomyces cerevisiae Oxidosqualene-Lanosterol Cyclase: A Chemistry-Biology Interdisciplinary Study of the Protein's Structure-Function-Reaction Mechanism Relationships
(2008)
Wu, Tung-Kung
;
Chang, Cheng-Hsiang
;
Liu, Yuan-Ting
;
Wang, Tsai-Ting
;
生物科技學系
The oxidosqualene cyclases (EC 5.4.99-) constitute a family of enzymes that catalyze diverse cyclization/rearrangement reactions of (3S)-2,3-oxidosqualene into a distinct array of sterols and triterpenes. The relationship between the cyclization mechanism and the enzymatic structure is extremely complex and compelling. This review covers the historical achievements of biomimetic studies and current progress in structural biology, molecular genetics, and bioinformatics studies to elucidate the mechanistic and structure-function relationships of the Saccharomyces cerevisiae oxidosqualene-lanosterol cyclase-catalyzed cyclization/rearrangement reaction. (C) 2008 The Japan Chemical journal Forum and Wiley Periodicals, Inc. Chem Rec 8: 302-325; 2008: Published online in Wiley, InterScience (www.interscience.wiley.com) DOI 10. 1002/tcr.20157
Site-saturated mutagenesis of histidine 234 of Saccharomyces cerevisiae oxidosqualene-lanosterol cyclase demonstrates dual functions in cyclization and rearrangement reactions
(2006-05-17)
Wu, Tung-Kung
;
Liu, Yuan-Ting
;
Chang, Cheng-Hsiang
;
Yu, Mei-Ting
;
Wang, Hisng-Ju
Site-saturated mutagenesis experiments were carried out on the His234 residue of Saccharomyces cerevisiae oxidosqualene-lanosterol cyclase (ERG7) to characterize its functional role in ERG7 activity and to determine its effect on the oxidosqualene cyclization/ rearrangement reaction. Two novel intermediates, (13 alpha H)-isomalabarica-14(26), 17E,21-trien-3 beta-ol and protosta-20,24-dien-3 beta-ol, isolated from ERG7(H234X) mutants, provided direct mechanistic evidence for formation of the chair-boat 6-6-5 tricyclic Markovnikov cation and protosteryl cation that were assigned provisionally to the ERG7-catalyzed biosynthetic pathway. In addition, we obtained mutants that showed a complete change in product specificity from lanosterol formation to either protosta-12,24-dien-3 beta-ol or parkeol production. Finally, the repeated observation of multiple abortive and/or alternative cyclization/ arrangement products from various ERG7(H234X) mutants demonstrated the catalytic plasticity of the enzyme. Specifically, subtle changes in the active site affect both the stability of the cation-pi interaction and generate product diversity.
The cysteine 703 to isoleucine or histidine mutation of the oxidosqualene-lanosterol cyclase from Saccharomyces cerevisiae generates an iridal-type triterpenoid
(2012-11-01)
Chang, Cheng-Hsiang
;
Chen, Yi-Chi
;
Tseng, Sheng-Wei
;
Liu, Yuan-Ting
;
Wen, Hao-Yu
The Cys703 to Ile or His mutation within Saccharomyces cerevisiae oxidosqualene-lanosterol cyclase ERG7 (ERG7(C7031/H)) generates an unusual truncated bicyclic rearranged intermediate, (8R,9R,10R)-polypoda-5,13E,17E,21-tetraen-3 beta-ol, related to iridal-skeleton triterpenoid. Numerous oxidosqualene-cyclized truncated intermediates, including tricyclic, unrearranged tetracyclic with 17 alpha/beta exocyclic hydrocarbon side chain, rearranged tetracyclic, and chair chair chair tricyclic intermediates (compounds 3-9), were also isolated from the ERG7(C703X) site-saturated mutations or the ERG7(F699T/C7031) double mutation, indicating the functional role of the Cys703 residue in stabilizing the bicyclic C-8 cation and the rearranged intermediate or interacting with Phe699, and opened a new avenue of engineering ERG7 for producing biological active agents. (C) 2012 Elsevier Masson SAS. All rights reserved.