• Title/Summary/Keyword: ARE-binding protein

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Primary Culture of Chicken Tracheal Epithelial Cells and Study on Those Characters for Recombinant Virus Infection (닭 기도 상피세포의 1차 배양과 유전자 재조합 바이러스의 감염 특성 연구)

  • Park, Mi Na;Jang, Hyun-Jun;Keum, Dae Ho;Choi, Jin Ae;Yoo, Jae Gyu;Byun, Sung June;Park, Jong Ju;Ji, Ju Young;Lee, Kyung-Tai;Kim, Tae-Hun;Lee, Hyun-Jeong
    • Korean Journal of Poultry Science
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    • v.40 no.4
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    • pp.299-304
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    • 2013
  • Tracheal epithelial cells (TECs) are an important tool for studies of viral respiratory diseases. Primary TECs have been cultured from human, mouse and hamster. It is also necessary to diagnose viral respiratory disease and reveal infection mechanisms in chicken. In this study, we isolated tracheal epithelial layers from tracheal of 20-day-old chicks and cultured primary TECs from the isolated layers. Ciliated cells which were a typical morphology of TECs were observed in cultured primary TECs and maintained until cell passage 5 (15 to 20 days). When we analyzed expression patterns of epithelial marker genes (retinoic acid responder, FGF-binding protein, virus activating protease (VAP) in TECs compared to immortalized chicken embryonic fibroblast cell line (DF-1), all the marker genes are highly expressed in TECs than in DF-1. When TECs were cultured with 0.1 and 1 MOI of ND virus (rNDV-GFP strain) to test the susceptibility of TECs for ND virus, 12.6% and 48.2% of the incubated TECs were infected respectively. In addition, when DF-1 was incubated with 1 MOI of ND virus, the virus infection rate of DF-1 was three times lower than the virus infection rate of TECs. These data could contribute to study infection mechanisms of viral respiratory diseases and control them in chicken.

Effects of High Glucose Levels on the Protein Kinase C Signal Transduction Pathway in Primary Cultured Renal Proximal Tubule Cells

  • Han, Ho-Jae;Kang, Ju-Won;Park, Kwon-Moo
    • The Korean Journal of Physiology
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    • v.30 no.2
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    • pp.257-267
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    • 1996
  • Diabetes mellitus is associated with a wide range of pathophysiologic changes in the kidney. This study was designed to examine the mechanisms by which glucose modulates the expression of polarized membrane transport functions in primary cultured rabbit renal proximal tubule cells. Results are as follows: The rate of 30 minute $Rb^{+}$ uptake was significantly higher($137.76{\pm}5.40%$) in primary renal tubular cell cultures treated with 20 mM glucose than that of 5 mM glucose. Not the level of mRNA for the ${\alpha}$ subunit of Na, K-ATPase but that of ${\beta}$ subunit was elevated in primary cultures treated with high glucose. The initial rate of methyl-${\alpha}$-D-glucopyranoside(${\alpha}$-MG) uptake was significantly lower($71.91{\pm}3.02%$) in monolayers treated with 20 mM glucose than that of 5 mM glucose. There was a tendency of an increase in phlorizin binding site in cells treated with 5 mM glucose. However, 3-O-methyl-D-glucose(3-O-MG) uptake was not affected by glucose concentration in culture media. TPA inhibited $Rb^{+}$ uptake by $63.61{\pm}1.94\;and\;45.80{\pm}1.36%$ and ${\alpha}$-MG uptake by $48.54{\pm}3.69\;and\;41.87{\pm}6.70%$ in the cells treated with 5 and 20 mM glucose, respectively. Also TPA inhibited mRNA expression of Na/glucose cotransporter in cells grown in 5mM glucose medium. cAMP significantly stimulated ${\alpha}$-MG uptake by $114.65{\pm}5.70%$ in cells treated with 5mM glucose, while it did not affect ${\alpha}$-MG uptake in cell treated with 20 mM glucose. However, cAMP inhibited $Rb^{+}$ uptake by $76.69{\pm}4.16\;and\;66.87{\pm}2.41%$ in cells treated with 5 and 20 mM glucose, respectively. In conclusion, the activity of the renal proximal tubular Na,K-ATPase is elevated in high glucose concentration. In contrast, the activity of the Na/glucose cotransport system is inhibited. High glucose may in part affect the activity of the Na,K-ATPase and the Na/glucose cotransport system by controlling the protein kinase C and/or A signal transduction pathway in primary cultured renal proximal tubule cells.

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Nutritive Evaluation of Some Browse Tree Legume Foliages Native to Semi-arid Areas in Western Tanzania

  • Rubanza, C.D.K.;Shem, M.N.;Otsyina, R.;Ichinohe, T.;Fujihara, T.
    • Asian-Australasian Journal of Animal Sciences
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    • v.16 no.10
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    • pp.1429-1437
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    • 2003
  • Browse tree legume leaves from Acacia spp (A. nilotica, A. tortilis, A. polyacantha), Dichrostachys sp, Flagea villosa, Piliostigma thonningii, Harrisonia sp were evaluated for nutritive potential (chemical compositions and degradability characteristics) compared to Gliricidia sepium. Effect of tannins anti-nutritive activity on digestibility was also assessed by polyethylene glycol (PEG) tannin bioassay. Crude protein (CP), ash, neutral detergent fiber (NDF), acid detergent fiber (ADF) and acid detergent lignin (ADL) differed (p<0.05) between legume foliages. Mean CP, ash, NDF, ADF and ADL for fodder species tested were 158, 92, 385, 145, and 100 g/kg DM, respectively. CP ranged from 115 (P. thonningii) to 205 g/kg DM (G. sepium). Acacia spp had moderate CP values (g/kg DM) of 144 (A. nilotica), to high CP in A. tortilis (188) and A. polyacantha (194) comparable to G. sepium. The forages had relatively lower fiber compositions. A. nilotica had (p<0.05) lowest NDF, ADF and ADL (182, 68 and 44) compared to P. thonningii (619, 196 and 130) g/kg DM, respectively. Except G. sepium, all fodder species had detectable high phenolic and tannin contents greater than 5% DM, an upper beneficial level in animal feeding and nutrition. Mean total phenolics (TP), total tannins (TT) and condensed tannins (CT) (or proanthocyanidins) for fodder species tested were 139, 113 and 43 mg/g DM, respectively. F. villosa had (p<0.05) lowest TP and TT of 65 and 56 mg/g DM, respectively, compared to A. nilotica (237 and 236 mg/g DM, respectively). The CT varied (p<0.05) from 6 (F. villosa) to 74 mg/g DM (Dichrostachys sp). In vitro organic matter (OM) degradability (OMD) differed (p<0.05) between fodder species. G. sepium had (p<0.05) high degradability potential compared to A. polyacantha that had (p<0.05) the lowest OMD values. Forage degradability ranked: G. sepium>A. nilotica>P. thonningi>F. villosa>Dichrostachys sp>A. tortilis>A. polyacantha. Addition of PEG resulted to (p<0.05) improvement in in vitro OM digestibility (IVD). Increase in IVD was mainly due to binding action of PEG on tannins; and represents potential nutritive values previously depressed by tannins anti-nutritive activity. Browse fodder has potential as sources of ruminal nitrogen especially for ruminants consuming low quality roughages due to high protein, lower fiber compositions and high potential digestibility. However, utilization of browse supplements in ruminants is hampered by high phenolic and tannin contents. Deactivation of tannin anti-nutritive activity, possibly by feeding tanniniferous browse with other readily available nitrogen sources to dilute tannin anti-nutritive activity could improve utilization of browse fodder supplements. Further studies are needed to assess browse fodder palatability and intake, and their effect on growth performance in ruminants.

Effect of Probiotics-Fermented Samjunghwan on Differentiation in 3T3-L1 Preadipocytes (3T3-L1 전지방세포에서 발효 삼정환의 지방 분화 억제 효과)

  • Song, Mi-Young;Bose, Shambhunath;Kim, Ho-Jun
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.42 no.1
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    • pp.1-7
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    • 2013
  • Samjunghwan (SJH) was fermented using five different probiotic bacterial strains (Lactobacillus plantarum, Enterococcus faecium, Pediococcus pentosaceus, Lactobacillus acidophilus or Bifidobacterium longum) separately. We examined the inhibition of preadipocyte differentiation through Oil Red O staining and analyzed the expression of CCAAT/enhancer-binding protein ${\alpha}$ ($C/EPB{\alpha}$), peroxisome proliferator-activated receptor ${\gamma}$ ($PPAR{\gamma}$), uncoupling protein (UCP)-2, and 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase which are adipogenic transcription factors. Both Lactobacillus plantarum and Enterococcus faecium-fermented SJH reduced Oil Red O dye staining compared with the same dose of non-fermented SJH. Only Lactobacillus plantarum-fermented SJH inhibited all adipogenic transcription factors and showed the best down-regulation of $PPAR{\gamma}$, UCP-2, and HMG-CoA reductase compared with the same dose of non-fermented SJH. The effect of SJH on the inhibition of preadipocyte differentiation was more prominent from the fermented SJH. Lactobacillus plantarum-fermented SJH, in particular, blocks the expression of $PPAR{\gamma}$, UCP-2, HMG-CoA reductase.

Molecular Cloning and Characterization of Trehalose Biosynthesis Genes from Hyperthermophilic Archaebacterium Metallosphaera hakonesis

  • Seo, Ju-Seok;An, Ju-Hee;Baik, Moo-Yeol;Park, Cheon-Seok;Cheong, Jong-Joo;Moon, Tae-Wha;Park, Kwan-Hwa;Choi, Yang-Do;Kim, Chung-Ho
    • Journal of Microbiology and Biotechnology
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    • v.17 no.1
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    • pp.123-129
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    • 2007
  • The trehalose $({\alpha}-D-glucopyranosyl-[1,1]-{\alpha}-D-glucopyranose)$ biosynthesis genes MhMTS and MhMTH, encoding a maltooligosyltrehalose synthase (MhMTS) and a maltooligosyltrehalose trehalohydrolase (MhMTH), respectively, have been cloned from the hyperthermophilic archaebacterium Metallosphaera hakonesis. The ORF of MhMTS is 2,142 bp long, and encodes 713 amino acid residues constituting a 83.8 kDa protein. MhMTH is 1,677 bp long, and encodes 558 amino acid residues constituting a 63.7 kDa protein. The deduced amino acid sequences of MhMTS and MhMTH contain four regions highly conserved for MTSs and three for MTHs that are known to constitute substrate-binding sites of starch-hydrolyzing enzymes. Recombinant proteins obtained by expressing the MhMTS and MhMTH genes in E. coli catalyzed a sequential reaction converting maltooligosaccharides to produce trehalose. Optimum pH of the MhMTS/MhMTH enzyme reaction was around 5.0 and optimum temperature was around 70 C. Trehalose-producing activity of the MhMTS/ MhMTH was notably stable, retaining 80% of the activity after preincubation of the enzyme mixture at $70^{\circ}C$ for 48 h, but was gradually abolished by incubating at above $85^{\circ}C$. Addition of thermostable $4-{\alpha}-glucanotransferase$ increased the yield of trehalose production from maltopentaose by 10%. The substrate specificity of the MhMTS/MhMTH-catalyzed reaction was extended to soluble starch, the most abundant maltodextrin in nature.

Polymorphisms and Functional Analysis of the Intact Human Papillomavirus16 E2 Gene

  • Ekalaksananan, Tipaya;Jungpol, Watcharapol;Prasitthimay, Chuthamas;Wongjampa, Weerayut;Kongyingyoes, Bunkerd;Pientong, Chamsai
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.23
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    • pp.10255-10262
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    • 2015
  • High risk human papillomavirus (HR-HPV) E2 proteins play roles in transcriptional regulation and are commonly functionally disrupted when the HPV genome integrates into host chromosomes. Some 15-40% of cancer cases, however, contain an intact E2 gene or episomal HPV. In these cases, polymorphism of the E2 gene might be involved. This study aimed to determine polymorphisms of the E2 gene in episomal HPV16 detected in high grade squamous intraepithelial lesions and squamous cell carcinomas and altered functions compared to the E2 prototype. The E2 gene was amplified and sequenced. Two expression vectors containing E2 gene polymorphisms were constructed and transfected in SiHa and C33A cells, then E6 gene as well as Il-10 and TNF-${\alpha}$ expression was determined by quantitative RT-PCR. Expression vectors and reporter vectors containing the HPV16 long control region (LCR) were co-transfected and transcriptional activity was determined. The results showed that a total of 32 nucleotides and 23 amino acids were changed in all 20 cases of study, found in the transactivation (TA) domain, hinge (H) region and DNA binding (DB) domain with 14, 5 and 13 nucleotide positions. They mostly caused amino acid change. The expressing vectors containing different E2 gene polymorphisms showed E6 mRNA suppression, TNF-${\alpha}$ mRNA suppression and IL-10 induction but no statistically significant differences when compared to the E2 prototype. Moreover, promoter activity in HPV16 LCR was not affected by E2 protein with different gene polymorphisms, in contrast to nucleotide variations in LCR that showed an effect on transcription activity. These results demonstrated that E2 gene polymorphisms of episomal HPV16 did not affect transcriptional regulation and suggested that nucleotide variation as well as epigenetic modification of the LCR might play a role in inducing malignant transformation of cells containing episomal HPV16.

Solid Phase Synthesis of Lysine-exposed Peptide-Polymer Hybrids by Atom Transfer Radical Polymerization (ATRP를 이용한 Lysine 말단기를 가진 펩타이드-고분자 하이브리드 합성)

  • Ha, Eun-Ju;Kim, Mijin;Kim, Jinku;An, Seong Soo A.;Paik, Hyun-Jong
    • Polymer(Korea)
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    • v.38 no.4
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    • pp.550-556
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    • 2014
  • Recently, the peptide(or protein)-polymer hybrid materials (PPs) were sought in many research areas as potential building blocks for assembling nanostructures in selective solvents. In PPs, the facile routes of preparing well-defined peptide-polymer bio-conjugates and their specific activities in various applications are important issues. Our strategy to prepare the peptide-polymer hybrid materials was to combine atom transfer radical polymerization (ATRP) method with solid phase peptide synthesis. The standard solid phase peptide synthesis method was employed to prepare the PYGK (proline-tyrosine-glycine-lysine) peptide. PYGK is an analogue peptide, PFGK (proline-phenylalanine-glycine-lysine), which interacted with plasminogen in fibrinolysis. The peptide and the peptide-initiator were characterized with MALDI-TOF mass spectrometry and $^1H$ NMR spectrometer. The peptide-polymer, pSt-PYGK was characterized by GPC, IR, $^1H$ NMR spectrometer and TLC. Spherical micellar aggregates were determined by TEM and SEM. Current synthesis methodology suggested opportunities to create the well-defined peptide-polymer hybrid materials with specific binding activity.

Chronic Treatment of Fluoxetine Increases Expression of NCAM140 in the Rat Hippocampus (장기간 플루세틴 처리에 의한 흰쥐 해마에서의 NCAM140 유전자 발현의 증가)

  • Choi, Mi Ran;Chai, Young Gyu;Jung, Kyoung Hwa;Baik, Seung Youn;Kim, Seok Hyeon;Roh, Sungwon;Choi, Joonho;Lee, Jun-Seok;Choi, Ihn Geun;Yang, Byung-Hwan
    • Korean Journal of Biological Psychiatry
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    • v.16 no.1
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    • pp.5-14
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    • 2009
  • Objectives : Most of the mechanisms reported for antidepressant drugs are the enhancement of neurite outgrowth and neuronal survival in the rat hippocampus. Neural cell adhesion molecule 140(NCAM140) has been implicated as having a role in cell-cell adhesion, neurite outgrowth, and synaptic plasticity. In this report, we have performed to elucidate a correlation among chronic antidepressant treatments, NCAM140 expression, and activation of phosphorylated cyclicAMP responsive element binding protein(pCREB) which is a downstream molecule of NCAM140-mediated intracellular signaling pathway in the rat hippocampus. Methods : Fluoxetine(10mg/kg) was injected acutely(daily injection for 5days) or chronically(daily injection for 14days) in adult rats. RNA and protein were extracted from the rat hippocampus, respectively. Real-time RT-PCR was performed to analyze the expression pattern of NCAM140 gene and western blot analyses for the activation of the phosphorylation ratio of CREB. Results : Chronic fluoxetine treatments increased NCAM140 expression 1.3 times higher than control in rat hippocampus. pCREB immunoreactivity in the rat hippocampus with chronic fluoxetine treatment was increased 4.0 times higher than that of control. Conclusion : Chronic fluoxetine treatment increased NCAM140 expression and pCREB activity in the rat hippocampus. Our data suggest that NCAM140 and pCREB may play a role in the clinical efficacy of antidepressants promoting the neurite outgrowth and neuronal survival.

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A Study on Iron Deficiency Anemia of Pre-School Children in Rural Area in Korea (한국농촌(韓國農材) 미취학아동(未就學兒童)의 철결핍성빈혈(鐵缺乏性貧血)에 관(關)한 연구(硏究) -충청남도(忠淸南道) 대덕군(大德郡) 유성면(儒城面) 상대리(上垈里)를 중심(中心)으로-)

  • Tchai, B.S.;Lee, Hyo-Eun
    • Journal of Nutrition and Health
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    • v.3 no.3
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    • pp.149-159
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    • 1970
  • This study is to investigate the status of anemia, especially iron deficiency anemia among pre-school children in rural area in Korea. The survey was conducted in Sang-dae Ri, Yusong Myon, Daedok Gun, Chung Chong Nam-Do from July 30 th to August 12th, 1968. The measurements were done of height, weight, hematologist and biochemical levels on ninety-two pre-school children, 47 male, and 45 female, one to six years of age. Hemoglobin was determined by the method of cyanmethemoglobin and hematocrit by micro hematocrit centrifuge. The determination of serum iron, iron-binding capacity was done by the method of Ramsay using bathophenanthroline and the serum albumin was determined by Biuret Reaction. The results of this study are as follows: 1) 54.4 percent of the pre-school children weighed less than 90 percent of the Korean General Standard Weight level. 2) The average hemoglobin level was $11.0{\pm}1.57gm/100ml$, 38.0 percent of the children were anemic with less than 1.0gm/100ml. Of the anemic children 60 percent were below the Korean General Standard Weight level. 3) 27.5 percent of the pre-school children were found to have below 32 percent of a hematocrit values and 28.0 percent showed less than 33 percent in M.C.H.C. These results showed that the incidence of hypochromic anemia in these pre-school children was high. 4) 37.9 percent of these children had a serum iron level less than $50{\mu}g/100ml\;and\;31.0\;percent\;had\;a\;TIBC\;above\;400{\mu}g$ while 48.3 percent showed a transferrin saturation lower than 15 percent. On the basis of these findings, it is concluded than the cause of this anemia was iron deficiency. 5) In this group there was a little evidence of low total serum protein levels. However, 10.4 percent of the children had a deficient serum albumin level, below 2.80 gm/100ml while 51.7 percent had a low level, less than 3.50gm/100ml, and 34.5 percent of the children had a low level of TIBC, less than $350{\mu}g/100ml$, and considering these facts, it is suggested that some of the anemias have a multiple causes through protein deficiency and repeated chronic infection apart from iron deficiency.

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Molecular Characterization of Extracellular Medium-chain-length Poly(3-hydroxyalkanoate) Depolymerase Genes from Pseudomonas alcaligenes Strains

  • Kim Do Young;Kim Hyun Chul;Kim Sun Young;Rhee Young Ha
    • Journal of Microbiology
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    • v.43 no.3
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    • pp.285-294
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    • 2005
  • A bacterial strain M4-7 capable of degrading various polyesters, such as poly$(\varepsilon-caprolactone)$, poly(3-hydroxybutyrate-co-3-hydroxyvalerate), poly(3-hydroxyoctanoate), and poly(3-hydroxy-5-phenylvalerate), was isolated from a marine environment and identified as Pseudomonas alcaligenes. The relative molecular mass of a purified extracellular medium-chain-length poly(3-hydroxyalkanoate) (MCL-PHA) depolymerase $(PhaZ_{palM4-7})$ from P. alcaligenes M4-7 was 28.0 kDa, as determined by SDS-PAGE. The $PhaZ_{palM4-7}$ was most active in 50 mM glycine-NaOH buffer (pH 9.0) at $35^{\circ}C$. It was insensitive to dithiothreitol, sodium azide, and iodoacetamide, but susceptible to p-hydroxymercuribenzoic acid, N-bromosuccinimide, acetic anhydride, EDTA, diisopropyl fluorophosphate, phenylmethylsulfonyl fluoride, Tween 80, and Triton X-100. In this study, the genes encoding MCL-PHA depolymerase were cloned, sequenced, and characterized from a soil bacterium, P. alcaligenes LB19 (Kim et al., 2002, Biomacro-molecules 3, 291-296) as well as P. alcaligenes M4-7. The structural gene $(phaZ_{palLB19})$ of MCL-PHA depolymerase of P. alcaligenes LB19 consisted of an 837 bp open reading frame (ORF) encoding a protein of 278 amino acids with a deduced $M_r$ of 30,188 Da. However, the MCL-PHA depolymerase gene $(phaZ_{palM4-7})$ of P. alcaligenes M4-7 was composed of an 834 bp ORF encoding a protein of 277 amino acids with a deduced Mr of 30,323 Da. Amino acid sequence analyses showed that, in the two different polypeptides, a substrate-binding domain and a catalytic domain are located in the N-terminus and in the C-terminus, respectively. The $PhaZ_{palLB19}$ and the $PhaZ_{palM4-7}$ commonly share the lipase box, GISSG, in their catalytic domains, and utilize $^{111}Asn$ and $^{110}Ser$ residues, respectively, as oxyanions that play an important role in transition-state stabilization of hydrolytic reactions.