• Title/Summary/Keyword: catalytic

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Studies on $\alpha$-amylase of Bocillus circulans F-2 (Part II) Enzymatic characteristics of the purified $\alpha$-amylase (Bacillus circulans F-2가 생산하는 $\alpha$-amylase에 관한 연구 (제 I I 보) 정제$\alpha$-amylase의 효소적특성)

  • ;Hajime Taniguchi;Yoshiharu Maruyama
    • Microbiology and Biotechnology Letters
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    • v.10 no.2
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    • pp.123-132
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    • 1982
  • These experiments were conducted to investigate the enzymatic characteristics of the purified $\alpha$-amylase (F-2A) of Bacillus circulans F-2 and the digestion rate of various starches. 1. The molecular weight was estimated to be 93000 by SDS-polyacrylamide disc gel electrophoresis. The isoelectric point was about pH 5.0. The optimum pH for the enzyme action was 6.0-6.5 and the stable pH ranged pH 5.5-12.0. The optimum temperature was 6$0^{\circ}C$, and the purified $\alpha$-amylase was stable below 4$0^{\circ}C$. 2. The purified $\alpha$-amylase was activated by Mn$^{++}$ and Co$^{++}$, whereas it was inhibited by Ag$^{+}$, HT$^{++}$, Cu$^{++}$ and Pb$^{++}$. 3. The purified $\alpha$-amylase is considered to have no sulfhydryl residue essential for its catalytic activity. 4. Michaelis constant (Km) was 1.704 mg/$m\ell$. Activation energy between 25-4$0^{\circ}C$ was 12.297 Kcal/mole, and between 40-6$0^{\circ}C$, it was 7.831 Kcal/mole. 5. The hydrolysis product from soluble starch, amylose and amylopectin in the early stage of hydrolysis was G$_{6}$, and as hydrolysis proceeds, G$_4$and G$_2$appeared. 6. Products from each oligosaccarides are as follows: G$_4$longrightarrow G$_2$+ G$_2$,G$_3$ +G$_1$,G$_{5}$longrightarrow G$_4$+G$_1$,G$_{6}$longrightarrowG$_4$+ G$_2$,G$_{7}$ G$_4$,G$_{8}$longrightarrow G$_4$+G$_4$, 7. On raw potato starch, raw sago starch and raw yam starch, the purified enzyme exhibited a remarkably high digestion rate than Porcine pancreatic amylase and Streptococcus bovis amylase.

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Role of Ascorbic Acid in the Depolymerization of Hyaluronic Acid by $Fe^{++}$ and $H_2O_2$ ($Fe^{++}$$H_2O_2$에 의한 hyaluronic acid 분해에 있어서 ascorbic acid의 역할)

  • Lee, Jung-Soo;Chung, Myung-Hee;Lim, Jung-Kyoo;Park, Chan-Woong;Cha, In-Joon
    • The Korean Journal of Pharmacology
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    • v.21 no.1
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    • pp.62-77
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    • 1985
  • In tile Iron-catalyzed Haber-Weiss reaction to produce OH., the requirement for $O^{-}_{2}{\cdot}$ is only to reduce $Fe^{+++}$. Possibly, the role of $O^{-}_{2}{\cdot}$ can be replaced by other reducing agents. Ascorbate is one of them in biological system. In the present study, the ability of ascorbate to produce $OH{\cdot}$ in the presence of $Fe^{++}$ and $H_2O_2$ was investigated by observing the degradation of hyaluronic acid and ethylene production from methional. Ascorbate stimulated the degradation of hyaluronic by $Fe^{++}$ and $H_2O_2$. That was confirmed by both viscosity change and gel-permeation chromatographic analysis. The observed degradation was almost completely prevented by catalase and $OH{\cdot}$ scavengers. In support of the above results, ascorbate enhanced the prouction of ethylene from methional in the presence of $Fe^{++}$ and $H_2O_2$. Other reducing agents (cysteine, glutathione, NADH and NADPH) showed similar activities to ascorbate in the degradation of hyaluronic acid and ethylene production. But no stimulatory effects were observed with their oxidized forms such as NAD and NADP. Thus, it appears that reduction of the metal ion was needed for $OH{\cdot}$ production. Among the metal ions tested, $Fe^{++}$ showed most potent catalytic action in the production of $OH{\cdot}$ The results obtained support that ascorbate can substitute $O^{-}_{2}{\cdot}$ in the metal-catalyzed reactions, particularly with $Fe^{++}$ by which $OH{\cdot}$ is produced with $H_2O_2$. The significance of the ascorbate-dependent production of $OH{\cdot}$ was considered with respect to possible role of ascorbate in the damage of inflamed joints.

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Transcriptomic Profile Analysis of Jeju Buckwheat using RNA-Seq Data (NA-Seq를 이용한 제주산 메밀의 발아초기 전사체 프로파일 분석)

  • Han, Song-I;Chung, Sung Jin;Oh, Dae-Ju;Jung, Yong-Hwan;Kim, Chan-Shick;Kim, Jae-hoon
    • Journal of the Korea Academia-Industrial cooperation Society
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    • v.19 no.1
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    • pp.537-545
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    • 2018
  • In this study, transcriptome analysis was conducted to collect various information from Fagopyrum esculentum and Fagopyrum tataricum during the early germination stage. Total RNA was extracted from the seeds and at 12, 24, and 36 hrs after germination of Jeju native Fagopyrum esculentum and Fagopyrum tataricum and sequenced using the Illumina Hiseq 2000 platform. Raw data analysis was conducted using the Dynamic Trim and Lengths ORT programs in the SolexaQA package, and assembly and annotation were performed. Based on RNA-seq raw data, we obtained 16.5 Gb and 16.2 Gb of transcriptome data corresponding to about 84.2% and 81.5% of raw data, respectively. De novo assembly and annotation revealed 43,494 representative transcripts corresponding to 47.5Mb. Among them, 23,165 sequences were shown to have similar sequences with annotation DB. Moreover, Gene Ontology (GO) analysis of buckwheat representative transcripts confirmed that the gene is involved in metabolic processes (49.49%) of biological processes, as well as cell function (46.12%) in metabolic process, and catalytic activity (80.43%) in molecular function In the case of gibberellin receptor GID1C, which is related to germination of seeds, the expression levels increased with time after germination in both F. esculentum and F. tataricum. The expression levels of gibberellin 20-oxidase 1 were increased within 12 hrs of gemination in F. esculentum but continuously until 36 hrs in F. tataricum. This buckwheat transcriptome profile analysis of the early germination stage will help to identify the mechanism causing functional and morphological differences between species.

Characteristics on De-CH4/NOx according to Ceramic and Metal Substrates of SCR Catalysts for CNG Buses (CNG 버스용 SCR 촉매의 세라믹과 메탈 담체에 따른 De-CH4/NOx 특성)

  • Seo, Choong-Kil
    • Journal of the Korea Academia-Industrial cooperation Society
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    • v.19 no.1
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    • pp.18-24
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    • 2018
  • The policy-making and technological development of eco-friendly automobiles designed to increase their supply is ongoing, but the internal combustion engine still accounts for about 95% of the automobiles in use. Also, in order to meet the stricter emission regulations of internal combustion engines based on fossil fuels, the proportion of after-treatments for vehicles and (ocean going) vessels is gradually increasing. Natural gas is a clean fuel that emits few air pollutants and has been used mainly as a fuel for city buses. In the long term, we intend to develop a new NGOC/LNT+NGCO/SCR combined system that simultaneously reduces the toxic gases, $CH_4$ and NOx, emitted from CNG buses. The objective of this study is to investigate the characteristics of $de-CH_4/NOx$ according to the ceramic and metal substrates of the SCR (Selective Catalytic Reduction) catalysts mounted downstream of the combined system. The V and Cu-SCR catalysts did not affect the $CH_4$ oxidation reaction, the two NGOC/SCR catalysts each coated with two layers began to oxidize $CH_4$ at $400^{\circ}C$, and the amount of $CH_4$ emitted was reduced to about 20% of its initial value at about $550^{\circ}C$. The two NGOC/SCR catalysts each coated with two layers showed a negative (-) NOx conversion rate above $350^{\circ}C$. The ceramic-based combined system reached LOT50 at $500^{\circ}C$, which was about 20% higher in terms of the $CH_4$ conversion rate than the metal-based combined system, showing that the combined system of NGOC/LNT+Cu-SCR is a suitable combination.

Iron chelating agent, deferoxamine, induced apoptosis in Saos-2 osteosarcoma cancer cells (Saos-2 골육종 세포에서 iron chelating agent, deferoxamine에 의한 apoptosis 유도)

  • Park, Eun Hye;Lee, Hyo Jung;Lee, Soo Yeon;Kim, Sun Young;Yi, Ho Keun;Lee, Dae Yeol;Hwang, Pyoung Han
    • Clinical and Experimental Pediatrics
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    • v.52 no.2
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    • pp.213-219
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    • 2009
  • Purpose:Iron is a critical nutritional element that is essential for a variety of important biological processes, including cell growth and differentiation, electron transfer reactions, and oxygen transport, activation, and detoxification. Iron is also required for neoplastic cell growth due to its catalytic effects on the formation of hydroxyl radicals, suppression of host defense cell activities, and promotion of cancer cell multiplication. Chronic transfusion-dependent patients receiving chemotherapy may have iron overload, which requires iron-chelating therapy. We performed this study to demonstrate whether the iron chelating agent deferoxamine induces apoptosis in Saos-2 osteosarcoma cells, and to investigate the underlying apoptotic mechanism. Methods:To analyze the apoptotic effects of an iron chelator, cultured Saos-2 cells were treated with deferoxamine. We analyzed cell survival by trypan blue and crystal violet analysis, apoptosis by nuclear condensation, DNA fragmentation, and cell cycle analysis, and the expression of apoptotic related proteins by Western immunoblot analysis. Results:Deferoxamine inhibited the growth of Saos-2 cell in a time- and dose-dependent manner. The major mechanism for growth inhibition with the deferoxamine treatment was by the induction of apoptosis, which was supported by nuclear staining, DNA fragmentation analysis, and flow cytometric analysis. Furthermore, bcl-2 expression decreased, while bax, caspase-3, caspase-9, and PARP expression increased in Saos-2 cells treated with deferoxamine. Conclusion:These results demonstrated that the iron chelating agent deferoxamine induced growth inhibition and mitochondrial-dependent apoptosis in osteosarcoma Saos-2 cells, suggesting that iron chelating agents used in controlling neoplastic cell fate can be potentially developed as an adjuvant agent enhancing the anti-tumor effect for the treatment of osteosarcoma.

Human Health Risk, Environmental and Economic Assessment Based on Multimedia Fugacity Model for Determination of Best Available Technology (BAT) for VOC Reduction in Industrial Complex (산업단지 VOC 저감 최적가용기법(BAT) 선정을 위한 다매체 거동모델 기반 인체위해성·환경성·경제성 평가)

  • Kim, Yelin;Rhee, Gahee;Heo, Sungku;Nam, Kijeon;Li, Qian;Yoo, ChangKyoo
    • Korean Chemical Engineering Research
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    • v.58 no.3
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    • pp.325-345
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    • 2020
  • Determination of Best available technology (BAT) was suggested to reduce volatile organic compounds (VOCs) in a petrochemical industrial complex, by conducting human health risk, environmental, and economic assessment based on multimedia fugacity model. Fate and distribution of benzene, toluene, ethylbenzene, and xylene (BTEX) was predicted by the multimedia fugacity model, which represent VOCs emitted from the industrial complex in U-city. Media-integrated human health risk assessment and sensitivity analysis were conducted to predict the human health risk of BTEX and identify the critical variable which has adverse effects on human health. Besides, the environmental and economic assessment was conducted to determine the BAT for VOCs reduction. It is concluded that BTEX highly remained in soil media (60%, 61%, 64% and 63%), and xylene has remained as the highest proportion of BTEX in each environment media. From the candidates of BAT, the absorption was excluded due to its high human health risk. Moreover, it is identified that the half-life and exposure coefficient of each exposure route are highly correlated with human health risk by sensitivity analysis. In last, considering environmental and economic assessment, the regenerative thermal oxidation, the regenerative catalytic oxidation, the bio-filtration, the UV oxidation, and the activated carbon adsorption were determined as BAT for reducing VOCs in the petrochemical industrial complex. The suggested BAT determination methodology based on the media-integrated approach can contribute to the application of BAT into the workplace to efficiently manage the discharge facilities and operate an integrated environmental management system.

Effect of Protein Kinase C Inhibitor (PKCI) on Radiation Sensitivity and c-fos Transcription Activity (Protein Kinase C Inhibitor (PKCI)에 의한 방사선 민감도 변화와 c-fos Proto-oncogene의 전사 조절)

  • Choi Eun Kyung;Chang Hyesook;Rhee Yun-Hee;Park Kun-Koo
    • Radiation Oncology Journal
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    • v.17 no.4
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    • pp.299-306
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    • 1999
  • Purpose : The human genetic disorder ataxia-telangiectasia (AT) is a multisystem disease characterized by extreme radiosensitivity. The recent identification of the gene mutated in AT, ATM, and the demonstration that it encodes a homologous domain of phosphatidylinositol 3-kinase (PI3-K), the catalytic subunit of an enzyme involved in transmitting signals from the cell surface to the nucleus, provide support for a role of this gene in signal transduction. Although ionizing radiation was known to induce c-fos transcription, nothing is known about how ATM or PKCI mediated signal transduction pathway modulates the c-fos gene transcription and gene expression. Here we have studied the effect of PKCI on radiation sensitivity and c-fos transcription in normal and AT cells. Materials and Methods: Normal (LM217) and AT (AT5BIVA) cells were transfected with PKCI expression plasmid and the overexpression and integration of PKCI was evaluated by northern blotting and polymerase chain reaction, respectively. 5 Gy of radiation was exposed to LM and AT cells transfected with PKCI expression plasmid and cells were harvested 48 hours after radiation and investigated apoptosis with TUNEL method. The c-fos transcription activity was studied by performing CAT assay of reporter gene after transfection of c-fos CAT plasmid into AT and LM cells. Results: Our results demonstrate for the first time a role of PKCI on the radiation sensitivity and c-fos expression in LM and AT cells. PKCI increased radiation induced apoptosis in LM cells but reduced apoptosis in AT cells. The basal c-fos transcription activity is 70 times lower in AT cells than that in LM cells. The c-fos transcription activity was repressed by overexpression of PKCI in LM cells but not in AT cells. After induction of c-fos by Ras protein, overexpression of PKCI repressed c-fos transcription in LM cells but not in AT cells Conclusion: Overexpression of PKCI increased radiation sensitivity and repressed c-fos transcription in LM cells but not in AT cells. The results may be a. reason of increased radiation sensitivity of AT cells. PKCI may be involved in an ionizing radiation induced signal transduction pathway responsible for radiation sensitivity and c-fos transcription. The data also provided evidence for novel transcriptional difference between LM and AT cells.

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Identification and Functional Analysis of Escherichia coli RNase E Mutants (Escherichia coli 리보핵산 내부분해효소 RNase E의 돌연변이체 선별 및 특성분석)

  • Shin, Eun-Kyoung;Go, Ha-Young;Kim, Young-Min;Ju, Se-Jin;Lee, Kang-Seok
    • Korean Journal of Microbiology
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    • v.43 no.4
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    • pp.325-330
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    • 2007
  • RNase E is an essential Escherichia coli endoribonuclease that plays a major role in the decay and processing of a large fraction of RNAs in the cell and expression of N-terminal domain consisted of 1-498 amino acids (N-Rne) is sufficient to support normal cellular growth. By utilizing these properties of RNase E, we developed a genetic system to screen for amino acid substitutions in the catalytic domain of the protein (N-Rne) that lead to various phenotypes. Using this system, we identified three kinds of mutants. A mutant N-Rne containing amino acid substitution in the S1 domain (I6T) of the protein was not able to support survival of E. coli cells, and another mutant N-Rne with amino acid substitution at the position 488 (R488C) in the small domain enabled N-Rne to have an elevated ribonucleolytic activity, while amino acid substitution in the DNase I domain (N305D) only enabled N-Rne to support survival of E. roli cells when the mutant N-Rne was over-expressed. Analysis of copy number of ColEl-type plasmid revealed that effects of amino acid substitution on the ability of N-Rne to support cellular growth stemmed from their differential effects on the ribonucleolytic activity of N-Rne in the cell. These results imply that the genetic system developed in this study can be used to isolate mutant RNase E with various phenotypes, which would help to unveil a functional role of each subdomain of the protein in the regulation of RNA stability in E. coli.

Studies on the Epidemiological Change of Clonorchiasis After Mass Chemotherapy in Highly endemic Areas (집단치료사업에 따른 간흡충증의 역학적 변동에 관한 조사)

  • Joo, Kyoung-Hwan;Chu, Paul B.;Rim, Han-Jong;Lee, Joon-Sang
    • Journal of agricultural medicine and community health
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    • v.12 no.1
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    • pp.80-93
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    • 1987
  • Mass chemotherapy of Clonorchis sinensis infection in Korea was started in 1982 with 40mg/kg body weight single dose administration scheme of praziquantel. The purpose of this study was to investigate the efficiency of current mass chemotherapy project and compare the epidemiological changes in endemic area of Ckmorchis sinensis. This study was performed at Kimhae-city and Samrangjin-eup of Miryang-gun, Kyongnam province, highly endemic area of C. sinensis located southeastern part of Korea from July to October of 1986. The therapy project of Kimhae area was performed in 1984, whereas that of Samrangjin was done in 1985 by stool examination of the Korea Association for Parasite Eradication(KAPE) and drug administration of local health centre. The results obtained were as follows; 1) As a results of stool examination from 234 specimens obtained in Kimhae area, the infection rate has decreased to 34.2%from 45.6p in 1983, but the infection rate of C. sinensis from 341 specimens obtained in Samrangjin area did not decrease (58.1%in 1986 490%in 1983). 2) The study in Kimhae area showed that the average EPG decreased remarkably from 4,858 to 1,340 and those classified above the category of heavy infection decreased also from 14.0pp to 1.7%. The study in Samrangjin area showed that the average EPG did decrease drastically from 9,597 to 6,498 and those classified above the category of heavy infection did not go down drastically from 25.2% to 14.2%. 3) The study in Kimhae area showed decrease of Cs.$D._{50}$ in comparison to that in 1983, wheareas Cs.$D._{50}$ in Samrangjin area showed no much difference compared to that in 1983. The intensities of endemicity were represented with the regression equation calculated with the cumulative percentages of EPG count. Regression equation was Y=4.49+1.19 log x in Kimhae area and Y=3.66+127 log x in Samrangjin area. 4) The two stage catalytic model was applied and the calculation lead to the equation $Y=5.33(e^{-0.018t}-e^{-0.016t})$ in Kimhae area and $Y=1.25(e^{-0.010t}-e^{-0.018t})$ in Kimhae area and $Y=125(e^{-0.010t}-e^{-0.050t})$ in Samrangjin area 5) The infection rate of cercaria in P.manchouric-us studied in Kimhae area showed 1.25% which is not much different from that in previous years, wheareas the infection rate of metacercaria in P. parva studied in the same area this year showed 2.5-20.2/gm of flesh in comparison to 64/gm of flesh in 1983. 6) Data of C. sinensis infection on the reservoir host in Kimhae area showed 4 out of 18 dogs, 1 out of 18 rats and that in Samrangjin area showed 2 out of 18 dogs respectively. 7) Among the inhabitants who were under mass chemotherapy in Kimhae area, 71out of them, upon stool examination, showed infection rate of 66.2% and those classified above the category of heavy infection, 2.4%. In comparison to infection rate of 33.7% and those classified above the category of heavy infection, which is 1.0%, obtained from those not under mass chemotherapy showed higher infection rate and somewhat equal distribution of intensity of infection. The above statements reflect the fact that individual therapy besides mass chemotherapy was prevalent in that area. 8) On the other side, the studies in Samrangjin area showed infection rate of 68.7% and those above the category of heavy infection, which is 6.1%, in comparison to infection rate of 58.3% and those above the category of heavy infection, which is 16.5%, in those not under mass chemotherapy. the above reflects that although a good deal of inhabit-ants were classified under light or moderate infection category, those above the category of heavy infection, yet, numbered a lot, and individual chemotherapy has not been going on. In conclusion, it was suggested that the number of reinfected inhabitants among those under mass chemotherapy were numerous. Accordingly, the reinforcement of health education should be followed with mass chemotherapy. The facts of high infection rate exemplified by 65% and high number of those above the category of heavy infection in Samrangjin area say that reevaluation of dosage, number of medication and intervals should be necessarily made.

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Biochemical Characterizations of Phenylalanine Ammonia-Lyase and its Mutants to Develop an Enzymatic Therapy for Phenylketonuria (페닐케톤뇨증의 효소치료 개발을 위한 phenylalanine ammonia-lyase 및 유전자 변이형의 생화학적 특성)

  • Kim, Woo-Mi
    • Journal of Life Science
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    • v.19 no.9
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    • pp.1226-1231
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    • 2009
  • Enzyme substitution with recombinant phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) is currently being explored for treatment of phenylketonuria (PKU), an autosomal recessive genetic disorder with mutations of the gene encoding phenylalanine-4-hydroxylase (EC 1.14.16.1). However, oral administration of PAL is limited because of proteolytic digestion in the gastrointestinal tract. The aim of this study was to determine the biochemical properties of PAL and delinate the susceptibility of wild-type PAL to pancreatic proteolysis by exploring several mutants, and to develop therapeutic drugs with PAL for PKU. The specific activity of PAL was assayed and its optimal pH, temperature stability, and intestinal protease susceptibility were investigated. Its $V_{max}$ values for phenylalanine and tyrosine were 1.77 and $0.47{\mu}mol$/ min/mg protein, respectively, and its $K_m$ values were $4.77{\times}10^{-4}$ and $4.37{\times}10^{-4}\;M$, respectively. PAL showed an optimal pH at 8.5, corresponding to the average pH range of the small intestine. It showed no loss of activity at $-80^{\circ}C$ for 5 months and possessed 93.4% of its activity under $4^{\circ}C$ for 4 wks. PAL was susceptible to chymotrypsin digestion and, to a lesser extent, to trypsin, elastase, carboxypeptidase A, and B. The trypsin and chymotrypsin cleaving sites were mutated to investigate protection from pancreatic digestion and the specific activities of these mutants were evaluated. The six mutants displayed low specific activities compared to the wild-type, suggesting that the primary trypsin and chymotrypsin cleaving sites may be essential for catalytic reaction. The PAL mutants could therefore be applied as a pretreatment modality without susceptibility to proteolytic attack, however, additional modification for enhancing enzymatic activity is needed to reduce the Phe levels effectively.