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Che-Chang Hsu

Publications and source records attributed to Che-Chang Hsu.

8 recordsLinked to original sources
PubMed2006

Release properties on gelatin-gum arabic microcapsules containing camphor oil with added polystyrene.

In this study, gelatin blended with arabic gum microcapsules containing camphor oil with added polystyrene were fabricated by a compound coacervation method. The parameters of oil/wall volume ratio, emulsification stirring speed, concentration of cross-linking agent, treated time and oil release properties were investigated. In order to improve the constant release effect of camphor oil, oil-soluble polystyrene (PS) was used as a sustained release agent. The camphor oil release curves were expressed by the exponential equation: psi(t)=C(eq)(1-e(-t/tau)), where psi(t) represent the variant of camphor oil concentration in the operation environment, C(eq) as the equilibrium state, t as the release time and tau as time constant. C(eq) and tau are significant factors pertaining to the camphor oil release properties. The results indicated that, for the microcapsules, the optimal oil/wall volume ratio was 0.75 to achieve the encapsulation efficiency of 99.6 wt.%. The average particle size were 294.7+/-14.2 microm, 167.2+/-11.2 microm, 85.7+/-8.7 microm at the homogenization stirring speed of 500, 1000, and 2000 rpm, respectively. The effect of sustained oil release will increase whereas the stirring speed decreases and the concentration of glutaraldehyde (GA) and treated time increases. Along with the increasing of quantity of polystyrene added, C(eq) decreased and tau increased, indicating that the sustained oil release amount and the release rate depend on the quantity of PS considerably.

Biocompatible Materials↗
PubMed2005

Directed evolution of D-sialic acid aldolase to L-3-deoxy-manno-2-octulosonic acid (L-KDO) aldolase.

An efficient L-3-deoxy-manno-2-octulosonic acid (L-KDO) aldolase was created by directed evolution from the Escherichia coli D-Neu5Ac (N-acetylneuraminic acid, D-sialic acid) aldolase. Five rounds of error-prone PCR and iterative screening were performed with sampling of 10(3) colonies per round. The specificity constant (kcat/Km) of the unnatural sugar L-KDO is improved to a level equivalent to the wild-type D-sialic acid aldolase for its natural substrate, D-Neu5Ac. The final evolved enzyme exhibits a >1,000-fold improved ratio of the specificity constant [kcat/Km (L-KDO)]/[kcat/Km (D-sialic acid)]. The protein sequence of the evolved aldolase showed eight amino acid changes from the native enzyme, with all of the observed changes occurring outside of the active site. Our effort demonstrates that an enzyme can be rapidly altered to accept enantiomeric substrates with screening of a small population of colonies iteratively toward the target substrate with improved catalytic efficiency. This work provides a method for the synthesis of enantiomeric sugars and for the study of enantiomeric catalysis affected by remote mutations.

Amino Acids↗
PubMed2003

One-pot synthesis of L-Fructose using coupled multienzyme systems based on rhamnulose-1-phosphate aldolase.

Two methods have been developed for the highly efficient enzymatic synthesis of L-fructose: one is based on rhamnulose-1-phosphate aldolase and acid phosphatase using racemic glyceraldehyde and dihydroxyacetone phosphate as substrates; the other is to generate enantiomerically pure L-glyceraldehyde in situ from glycerol for the aldol reaction, using galactose oxidase catalyzed oxidation of glycerol in the presence of catalase. Using this four-enzyme system, enantiomerically pure L-fructose was obtained. Using the more expensive dihydroxyacetone phosphate, the yield was 55% after purification.

Acid Phosphatase↗
PubMed2003

Directed evolution of N-acetylneuraminic acid aldolase to catalyze enantiomeric aldol reactions.

Expanding the scope of substrate specificity and stereoselectivity is of current interest in enzyme catalysis. Using error-prone PCR for in vitro directed evolution, the Neu5Ac aldolase from Escherichia coli has been altered to improve its catalytic activity toward enantiomeric substrates including N-acetyl-L-mannosamine and L-arabinose to produce L-sialic acid and L-KDO, the mirror-image sugars of the corresponding naturally occurring D-sugars. The first generation variant containing two mutations (Tyr98His and Phe115Leu) outside the (alpha,beta)(8)-barrel active site exhibits an inversion of enantioselectivity toward KDO and the second generation variant contains an additional amino acid change Val251Ile outside the alpha,beta-barrel active site that improves the enantiomeric formation of L-sialic acid and L-KDO. The X-ray structure of the triple mutant epNanA.2.5 at 2.3A resolution showed no significant difference between the wild-type and the mutant enzymes. We probed the potential structural 'hot spot' of enantioselectivity with saturation mutagenesis at Val251, the mutated residue most proximal to the Schiff base forming Lys165. The selected variant had an increase in k(cat) via replacement with another hydrophobic residue, leucine. Further sampling of a larger sequence space with error-prone PCR selected a third generation variant with significant improvement in L-KDO catalysis and a complete reversal of enantioselectivity.

Aldehydes↗
PubMed2003

Demonstration of functional coupling between gamma -aminobutyric acid (GABA) synthesis and vesicular GABA transport into synaptic vesicles.

l-Glutamic acid decarboxylase (GAD) exists as both membrane-associated and soluble forms in the mammalian brain. Here, we propose that there is a functional and structural coupling between the synthesis of gamma-aminobutyric acid (GABA) by membrane-associated GAD and its packaging into synaptic vesicles (SVs) by vesicular GABA transporter (VGAT). This notion is supported by the following observations. First, newly synthesized [(3)H]GABA from [(3)H]l-glutamate by membrane-associated GAD is taken up preferentially over preexisting GABA by using immunoaffinity-purified GABAergic SVs. Second, the activity of SV-associated GAD and VGAT seems to be coupled because inhibition of GAD also decreases VGAT activity. Third, VGAT and SV-associated Ca(2+)calmodulin-dependent kinase II have been found to form a protein complex with GAD. A model is also proposed to link the neuronal stimulation to enhanced synthesis and packaging of GABA into SVs.

Animals↗