• Journal of Sericultural and Entomological Science
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• v.39 no.2
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• pp.169-173
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• 1997

The behavior of tyrosine(Tyr.) residue of Bombyx mori silk fiber from yellow fluorescent cocoon has been examined for the dependence of pH in aqueous silk solution under the presence of orange II salt. Through the peak separation of angular dependence of spectral pattern of 15N-Tyr. and [1-13C]-Tyr. between the fiber axis and the molecular bond direction, N-H bond in fiber as well as the orientation distribution around the fiber axis were analyzed. Also, and sericin component was obtained from these angular dependence of oriented spectral pattern. The pH dependence of the 13C NMR chemical shift of B. mori silk fibroin was examined in aqueous solution in the presince of orange II are broad at pH$\geq$7.0. However, these become sharper at pH$\geq$8.0 and remain sharp at higher pH. In these higher pH range, a chemical shift change occurs due to the deprotonation of the Tyr. side group of fibroin. At higher pH. such a hydrophobic cluster is destroyed because of the electrostatic interaction according to the deprotonation of the Tyr-OH group.

• Journal of Life Science
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• v.9 no.2
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• pp.44-48
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• 1999

The ionization of tyrosine residue is known to be involved in the stabilization of transition-state in catalysis of astacin based upon the astacin-transition state analog structure. Two tyrosine residues, Tyr-216 and Tyr-219, are conserved in all MMPs related with astacin family, We replaced Tyr-216 and Tyr-219 into phenylalanine, respectively and the zinc binding properties, kinetic parameters, and pH dependence of each mutant are determined in order to examine the role of tyrosine residue in matrilysin catalysis. Both mutants contain two zinc atoms per mol of enzyme, indicating that either tyrosime does not affect the zinc binding property of the enzyme. Y216F and Y219F mutants are highly active and the kcat/Km values are only decreased 1.1-1.5-fold compared to the wild-type enzyme. The decrease in the activity of the mutants is essentially due to the increase in Km value. The pH dependencies of the kcat/Km values for both mutants are similar to the corresponding dependencies obtained with the wild type enzyme. The pKa values at the alkaline side of both mutants are not changed. These kinetic and pH dependence results indicate that the ionization of active site tyrosine residue of matrilysin is not reflected in the kinetics of peptide hydrolysin as catalyzed by astacin.

• KSBB Journal
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• v.7 no.3
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• pp.201-208
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• 1992

Cell surface binding of epidermal growth factor(EGF) to EGF receptors was studied for a series of site-directed receptor mutants transfected into B82 mouse fibroblasts. Scatchard plots for truncation mutant receptors significantly lost nonlinearity for truncations below residue 1022. Transient plots of dissociation kinetics exhibited biphasic behavior for all receptor types, but the fraction of receptor in slow-dissociating form was reduced by an order of magnitude for the truncation mutants below residue 1022. Comparison of dissociation kinetics between control cells and cells treated with Triton X-100 revealed no significant variation for the slow-dissociating receptor form, but a noticeable variation was observed for the fast-dissociating receptor form when EGF receptors were truncated below residue 991. These results suggest that high affinity of EGF binding at cell surface depend on the EGF receptor cytoplasmic region.

• Bulletin of the Korean Chemical Society
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• v.28 no.2
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• pp.271-275
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• 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.

• Mass Spectrometry Letters
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• v.10 no.2
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• pp.50-55
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• 2019

The fragmentation statistics of ion trap CID (Collision-Induced Dissociation) spectra using 87,661 tandem mass spectra of doubly charged tryptic peptides are analyzed here. In contrast to the usual method of using intensity information, the frequency of occurrence of fragment ions, with respect to the position of the cleavage site and the residues at these sites is studied in this paper. The analysis shows that the frequency of occurrence of fragment ion peaks is more towards the middle of the peptide than its ends. It was noted that amino acid with an aromatic and basic side chain at N- & C- terminal end of the peptide stimulates more peaks at the lower end of the spectrum. The residue pair effect was shown when the amide bond occurs between acidic and basic residues. The fragmentation at these sites (D/E-H/R/K) stimulates the generation of the y-ion peak. Also, the cleavage site H-H/R/K stimulates the generation of b-ions. K-P environment in the peptide sequence has more tendency to generate y-ions than b-ions. Statistical analysis helps in the visualization of the CID fragmentation pattern. Cleavage pattern along the length of the peptide and the residue pair effects, enhance the knowledge of fragmentation behavior, which is useful for the better interpretation of tandem mass spectra.

• Bulletin of the Korean Chemical Society
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• v.21 no.4
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• pp.379-382
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• 2000

The temperature dependence of the kinetic parameters of the aspartase-catalyzed reaction has been examined in the direction of deamination. The pK1values at 37$^{\circ}C$, 25$^{\circ}C$, 16$^{\circ}C$ and 7$^{\circ}C$ were 6.2 $\pm$ 0.1, 6.3 $\pm$ 0.1, $6.7{\pm}0.3$ and 6.9 $\pm$ 0.3, respectively. On the other hand, the pK2 values at 37$^{\circ}C$,25$^{\circ}C$, 16$^{\circ}C$ and 7$^{\circ}C$ were 8.1 $\pm0.2$, 8.3 $\pm$ 0.2, 8.2 $\pm$ 0.3 and 8.0 $\pm$ 0.2,respectively. The enthalpy of ionization, DHion, calculated from the slope of pK1, are 6.0 $\pm$ 0.3 kcal/mol. These results validate the prediction that aspartase requires a histidine residue for a general base, and a cysteine (or having a carboxyl functional group) for a general acid.

• BMB Reports
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• v.32 no.4
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• pp.326-330
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• 1999

An aspartase purified from Hafnia alvei was inactivated by diethylpyrocarbonate (DEP) in a pseudo-first-order inactivation. The first-order plot was biphasic. The inactivation process was not saturable and the second order rate constant was $1.3\;M^{-1}s^{-1}$. The inactivated aspartase was reactivated with NH₂OH. The difference absorption spectrum of DEP-inactivated vs native enzyme preparations revealed a marked peak around 242 nm. The pH dependence of the inactivation rate suggests that an amino acid residue having a pK value of 7.2 was involved in the inactivation. L-aspartate, fumarate (substrates), and chloride ion (inhibitor) protected the enzyme against inactivation, indicating that histidine residues for the enzyme activity are located at the active site of this aspartase. Inspection of the presence and absence of $Cl^-$ ion demonstrated that the number of essential histidine residues is less than two. Thus, one or two histidines are in or near the aspartate binding site and participate in an essential step of the catalytic reaction.

• BMB Reports
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• v.30 no.2
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• pp.113-118
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• 1997

Aspartase from Hafnia alvei was inactivated by N-ethylmaleimide (NEM) and 5,5' -Dithiobis-(2-znitrobenzoic acid) (DTNB) following pseudo-first order kinetics. Their apparent reaction orders were 0.83 and 0.50 for NEM and DTNB modifications, respectively, indicating that inactivation was due to a sulfhydryl group in the active site of aspartase and participation of the sulfhydryl group in an essential step in the catalytic reaction. When aspartase was modified by DTNB, the enzyme activity was restored by dithiothreitol treatment, indicating that cysteine residuetsl islarel possibly at or near the active site. The pH-dependence of the inactivation rate by NEM suggested that an amino acid residue having pK value of 8.3 was involved in the inactivation. When aspartase was incubated with NEM and L-aspartate together, L-aspartate markedly protected the enzyme from inactivation by NEM, but the other reagents used did not.

• Proceeding of EDISON Challenge
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• 2014.03a
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• pp.225-235
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• 2014

6종의 Intermediate filament 중 type IV인 Neurofilaments (NFs)는 신경세포에 존재하는 세포골격세사로 heavy NF(NF-H), medium NF(NF-M), light NF(NF-L) 세가지의 분자 질량 단백질로 구성되어 있다. NF의 side arm은 interfilament spacing과 axonal caliber를 조절하는 중요한 역할을 한다고 생각되어왔다. 또한 이에 대해서 각각의 protein의 역할은 알아내기 위해 isolated NF의 형태와 구조에 대해 많은 연구가 이루어졌는데, NF의 구조적 특성은 NF sidearm의 tail 부분에서 phosphorylation의 정도에 따른 Lys-Ser-Pro(KSP) repeats의 charge distribution을 통해 알 수 있다. 지금까지 NF에 대한 많은 연구가 이루어졌지만 인간에 한해서만 진행되었다. 그렇기 때문에 본 연구에서는 주어진 amino acid sequence와 각 species의 NF-H:NF-M:NF-L의 비율의 정보를 이용하여 The constant-NVT ensemble MC simulation을 통해 인간뿐만이 아닌 다른 species에 대한 NF의 구조적 특성을 알아보고자 한다.

• Journal of Life Science
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• v.12 no.1
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• pp.32-42
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• 2002

It was researched to react horse serum-BChE with hefanoylthiocholine chosen among choline esters. According as number of carbon of acyl group in choline esters was increased, reactivity was decreased but strength of ES complex was increased (Km=0,140mM). The pH-V/K profile for BChE-catalyzed hydrolysis of hexanoylthiocholine yields a p $K_{a}$ =4.974$\pm$0.028. This value is equal to recent literature that shows systematic shift from dependence of activity on the basic form fo a residue that huts a p $K_{a}$ =6.2~6.4 to catalysis by a residue or residues that has a p $K_{a}$ =4.7~5.0. The resulting kinetic solvent isotope effect of hexanoylthiocholine is $^{D/V}$K=1.18. The magnitude of the isotope effect suggests that proton transfer is not an element of transition-state stabilization.n.