• Journal of Microbiology and Biotechnology
• /
• 제2권1호
• /
• pp.56-65
• /
• 1992

The complete nucleotide sequence of the Bacillus circulans F-2 RSDA gene, coding for raw starch digesting a-amylase (RSDA), has been determined. The RSDA structure gene consists of an open reading frame of 2508 bp. Six bp upstream of the translational start codon of the RSDA is a typical gram-positive Shine-Dalgarno sequence and the RSDA encodes a preprotein of 836 amino acids with an Mr of 96, 727. The gene was expressed from its own regulatory region in E. coli and two putative consensus promoter sequences were identified upstream of a ribosome binding site and an ATG start codon. Confirmation of the nucleotide sequence was obtained and the signal peptide cleavage site was identified by comparing the predicted amino acid sequence with that derived by N-terminal analysis of the purified RSDA. The deduced N-terminal region of the RSDA conforms to the general pattern for the signal peptides of secreted prokaryotic proteins. The complete amino acid sequence was deduced and homology with other enzymes was compared. The results suggested that the Thr-Ser-rich hinge region and the non-catalytic domain are necessary for efficient adsorption onto raw substrates, and the catalytic domain (60 kDa) is necessary for the hydrolysis of substrates, as suggested in previous studies (8, 9).

• Journal of Microbiology and Biotechnology
• /
• 제28권8호
• /
• pp.1339-1345
• /
• 2018

2-Keto-3-deoxy-6-phosphogluconate (KDPG) aldolase, which catalyzes aldol cleavage and condensation reactions, has two distinct substrate-binding sites. The substrate-binding mode at the catalytic site and Schiff-base formation have been well studied. However, structural information on the phosphate-binding loop (P-loop) is limited. Zymomonas mobilis KDPG aldolase is one of the aldolases with a wide substrate spectrum. Its structure in complex with the substrate-mimicking 3-phosphoglycerate (3PG) shows that the phosphate moiety of 3PG interacts with the P-loop and a nearby conserved serine residue. 3PG-binding to the P-loop replaces water molecules aligned from the P-loop to the catalytic site, as observed in the apostructure. The extra electron density near the P-loop and comparison with other aldolases suggest the diversity and flexibility of the serine-containing loop among KDPG aldolases. These structural data may help to understand the substrate-binding mode and the broad substrate specificity of the Zymomonas KDPG aldolase.

• Bulletin of the Korean Chemical Society
• /
• 제28권2호
• /
• pp.271-275
• /
• 2007

γ -Glutamyl transpeptidase (EC 2.3.2.2) plays a key role in glutathione metabolism by catalyzing the transfer of the γ -glutamyl residue and hydrolysis of glutathione. The functional residues at the active site of the rat kidney γ -glutamyl transpeptidase were investigated by kinetic studies at various pH, the treatment of diethylpyrocarbonate (DEPC), and photooxidation in presence of methylene blue. An ionizable group affecting the enzymatic activity with an apparent pKa value of 7.1, which is in the range of pKa values for a histidine residue in protein, was obtained by examining the pH-dependence of kinetic parameters. The pH effect on the photoinduced inactivation rate of the enzyme corresponds to that expected for the photooxidation of the free histidine. The involvement of a histidine in the catalytic site of the enzyme was further supported by DEPC modification accompanied by an increase in absorbance at 240 nm, indicating the formation of Ncarbethoxyhistidine. The histidine located at the position of 382 in the precursor of the enzyme is primarily suspected based on the amino acid sequence alignment of the transpeptidases from various organisms.

• Bulletin of the Korean Chemical Society
• /
• 제22권1호
• /
• pp.77-83
• /
• 2001

To gain further insight into the relationship between structure and function of glutathione S-transferase (GST), the four cysteine mutants, C14S, C47S, C101S and C169S, of human GST P1-1 were expressed in Escherichia coli and purified to electrophoretic homogeneity by affinity chromatography on immobilized glutathione (GSH). The catalytic activities of the four mutant enzymes were characterized with five different substrates as well as by their binding to four different inhibitors. Cys14 seems to participate in the catalytic reaction of GST by stabilizing the conformation of the active-site loop, not in the GSH binding directly. The substitution of Cys47 with serine significantly reduces the affinity of GSH binding, although it does not prevent GSH binding. On the other hand, the substitution of Cys101 with serine appears to change the binding affinity of electrophilic substrate by inducing a conformational change of the $\alpha-helix$ D. Cys169 seems to be important for maintaining the stable conformation of the enzyme. In addition, all four cysteine residues are not needed for the steroid isomerase activity of human glutathione S-transferase P1-1.

• 공업화학
• /
• 제3권4호
• /
• pp.663-669
• /
• 1992

분말의 $Zr(OH)_4$를 $(NH_4)_2CrO_4$수용액에 함침시킨 후 공기중에 소성하여 질코니아에 담지된 산화크롬 촉매를 제조하였다. 제조된 $CrO_x/ZrO_2$ 촉매상에 cumene을 반응시켜 산화-환원 거동을 연구한 결과 크롬의 산화상태가 +6인 chromate형태로 질코니아 표면에 존재할 때는 강한 산점이 생성되어 cumene의 탈알킬화 반응에 촉매활성을 나타내었다. 그러나 많은 양의 $Cr^{6+}$종은 반응중에 탈수소로 생성된 $H_2$에 의하여 환원되어 $Cr^{3+}$종으로 변환되었으며 $Cr^{3+}$종은 cumene을 탈수소화시켜 ${\alpha}$-methyl styrene을 생성하는 반응에 활성점으로 작용하였다. 환원된 $Cr^{3+}$종은 $O_2$로 처리하면 다시 $Cr^{6+}$종으로 되고 따라서 산화된 촉매는 cumene의 탈알킬화 반응에 활성을 나타내었다.

• 한국진공학회:학술대회논문집
• /
• 한국진공학회 2013년도 제45회 하계 정기학술대회 초록집
• /
• pp.142.2-142.2
• /
• 2013

The effect of substrate on catalytic activity of CO oxidation with transition metal Platinum nanoparticles on doped and undoped TiO2 was investigated. Titanium dioxide was doped chemically with non-metal anions including nitrogen and fluorine. Undoped TiO2 was synthesized via simple conventional sol-gel route. Thin films of titania were developed by spin coating technique and the characterization techniques SEM, XRD, UV-Vis Absorption Spectroscopy and XPS were carried out to examine the morphology of films, crystal phase, crystallites, optical properties and elemental composition respectively. XPS analysis from doped TiO2 confirmed that the nitrogen site were interstitial whereas fluorine was doped into TiO2 lattice substitutionally. Catalytic activity systems of Pt/doped-TiO2 and Pt/undoped-TiO2 were fabricated to reveal the strong metal-support interaction effect during catalytic activity of CO oxidation reactions. By arc plasma deposition technique, platinum nanoparticles with mean size of 2.7 nm were deposited on the thin films of doped and undoped titanium dioxide. The CO oxidation was performed with 40 Torr CO and 100 Torr O2 with 620 Torr He carrier gas. Turn over frequency was observed two to three folds enhancement in case of Pt/doped TiO2 as compared to Pt/TiO2. The electronic excitation and the oxygen vacancies that were formed with the doping process were the plausible reasons for the enhancement of catalytic activity.

• 한국수소및신에너지학회논문집
• /
• 제33권1호
• /
• pp.105-112
• /
• 2022

Copper (Cu)-based catalysts have been widely used in a methanol steam reforming (MSR) reaction for hydrogen production for air-independent propulsion (AIP) applications and their good catalytic activities have attracted much attention. However, the agglomeration of the catalytic active site Cu causes deteriorating the catalytic performance and suppression of Cu agglomeration is a crucial issue in the AIP applications that the MSR system is typically operated at 250-300℃ for a long time. R. Sakai et al. recently showed a computational study on the anchoring effect that reduces an agglomeration of active sites by doping in a supporter. In order to present the anchoring effect on 𝛾-Al₂O₃ supported Cu-based catalysts, in this study, the adsorption energies of Cu on X-doped (X=ruthenium, phosphorus, silicon) 𝛾-Al₂O₃ were calculated and Cu adsorption energy decreased due to a change of the electronic structure originated from doping, thereby proving the anchoring effect.

• Journal of Microbiology and Biotechnology
• /
• 제30권7호
• /
• pp.1104-1107
• /
• 2020

The N-terminal domain of the Pseudomonas sp. FB15 phytase increases low-temperature activity and catalytic efficiency. In this study, the 3D structure of the N-terminal domain was predicted and substitutions for the amino acid residues of the region assumed to be the active site were made. The activity of mutants, in which alanine (A) was substituted for the original residue, was investigated at various temperatures and pH values. Significant differences in enzymatic activity were observed only in mutant E263A, suggesting that the amino acid residue at position 263 of the N-terminal domain is important in enzyme activity.

• BMB Reports
• /
• 제29권1호
• /
• pp.17-21
• /
• 1996

Ile91 of restriction endonuclease EcoRV, which has not been known to take part directly in catalytic activity, was substituted with Leu by site-directed mutagenesis. The Ile91Leu mutant shows over 1000-fold less activity than the wild type EcoRV under standard reaction condition. The metal ion dependency of the reaction was altered. In contrast to the wild type EcoRV, the mutant prefers $Mn^{2+}$ to $Mn^{2+}$ as the cofactor. In $Mn^{2+}$ buffer the mutant is as active as the wild type enzyme in $Mn^{2+}$ buffer. Like the wild type enzyme, the mutant shows an unspecific binding of DNA in gel shift experiments. In contrast to the wild type enzyme, the mutant did not cleave at noncognate sites of DNA under star condition.

• Archives of Pharmacal Research
• /
• 제20권2호
• /
• pp.115-121
• /
• 1997

Ile-269 in horse liver alcohol dehydrogenase isoenzyme S(HLADH-S) was mutated to serine by phosphorothioate-based site-directed mutagenesis in order to study the role of the residue in coenzyme binding. The specific activity of the mutant(1269S) enzyme to ethanol was increased 49-fold. All turnover numbers of 1269S enzyme toward 9 primary alcohols were increased. The mutant enzyme showed 3.6, 4.6, 11.6-fold higher catalytic efficiency for $5{\beta}$-androstane-3, 17-dione, $5{\beta}$-cholanic acid-3-one and retinal than wild-type, respectively. The reaction mechanism of 1269S enzyme was ordered bi bi as wild-type's. These results indicate that the hydrophobic interaction of Ile-269 residue with coenzyme plays an important role in dissociation of coenzyme from enzyme-coenzyme complex, which has been known as the rate limiting step of ADH reaction.