• Journal of Life Science
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• v.17 no.6 s.86
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• pp.772-777
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• 2007

A rice OsLeuD gene for small subunit of 3-isopropylmalate isomerase (IPMI) (EC 4.2.1.33) has been isolated. OsLeuD gene is located on 109.3 cM of chromosome 2. OsLeuD gene was expressed abundantly in metabolically active organs including leaves and developing seeds, indicating that OsLeuD gene expression is developmentally regulated. The cDNA of OsLeuD gene was coded for 257 amino acids which showed 58% and 48% homology to small subunits of IPMI in OsLeuD genes of cyanobacteria and green sulfur bacteria, respectively. The molecular character of OsLeuD is closely related to those of photosynthetic bacteria rather than those of eukaryotes including fungi and yeast. This suggests that OsLeuD gene in chromosomal genome of plants may possibly be originated from chloroplast genome.

• Applied Biological Chemistry
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• v.34 no.4
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• pp.334-338
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• 1991

Synthesis of $(D-Phe^7\;-Leu^8)$ bradykinin and bradykinin by solid phase method using a new reaction vessel was carried out. Coupling was performed by dicyclohexylcarbodiimide. After cleavage with dried HBr the peptides were purified by high pressure liquied chromatography. Their purify was assayed by paper and thin layer chromatography, melting point and amino acid analysis. $(D-Phe^7\;-Leu^8)$ bradykinin and bradykinin were incubater in vitro endopeptidase $({\alpha}-chymotrysis)$ and exopeptidase(carboxypeptidase A, leucine aminopeptidase) in order to study the degradation pattern of peptides. $(D-Phe^7\;-Leu^8)$ bradykinin and bradykinin were rapidly degradated by ${\alpha}-chymotrypsin$ and carboxypeptidase A $(D-Phe^7\;-Leu^8)$ bradykinin and bradykinin coution$(D-Phe^7\;-Leu^8)$ bradykinin and bradykinin contain imino peptide bound from proline at N-terminal and therefore they were not attacted by leucine aminopeptidase.

• Bulletin of the Korean Chemical Society
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• v.31 no.4
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• pp.874-880
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• 2010

A series of 13- and 14-membered cyclic enkephalin analogs based on the moderately $\mu$ selective prototype compound Tyr-C[D-$A_2bu$-Gly-Phe-Leu] 8a were synthesized to investigate the structure-activity relationship. The modifications of sequence were mainly focused on two positions 3 and 5, critical for the selective recognition for $\mu$ and $\delta$ opioid receptors. The substitution of hydrophobic $Leu^5$ with hydrophilic $Asp^5$ derivatives led to Tyr-C[D-$A_2bu$-Gly-Phe-Asp(N-Me)] 7 and Tyr-C[D-Glu-Phe-gPhe-rAsp(O-Me)] 5, the peptides with a large affinity losses at both $\mu$ and $\delta$ receptors. The substitution of $Phe^3$ with $Gly^3$ led to Tyr-C[D-Glu-Gly-gPhe-rLeu] 3 and Tyr-C[D-Glu-Gly-gPhe-D-rLeu] 4, the peptides with large affinity losses at $\mu$ receptors, indicating the critical role of phenyl ring of $Phe^3$ for $\mu$ receptor affinities. One atom reduction of the ring size from 14-membered analogs Tyr-C[D-Glu-Phe-gPhe-(L and D)-rLeu] 6a, 6b to 13-membered analogs Tyr-C[D-Asp-Phe-gPhe-(L and D)-rLeu] 1, 2 reduced the affinity at both $\mu$ and $\delta$ receptors, but increased the potency in the nociceptive assay, indicating the ring constrain is attributed to high nociceptive potency of the analogs. For the influence of D- or L-chirality of $Leu^5$ on the receptor selectivity, regardless of chirality and ring size, all cyclic diastereomers displayed marked $\mu$ selectivity with low potencies at the $\delta$ receptor. The retro-inverso analogs display similar or more active at $\mu$ receptor, but less active at $\delta$ receptor than the parent analogs.

• YAKHAK HOEJI
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• v.38 no.5
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• pp.530-543
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• 1994

To study the feasibility of transmucosal delivery of leucine enkephalin (Leu-Enk) and $[D-ala^2]$-leucine enkephalinamide (YAGFL), their degradation extents and pathways in various rabbit mucosa extracts were investigated by high performance liquid chromatography. The degradation of Leu-Enk and YAGFL was observed to follow the first-order kinetics. The degradation half-lives of Leu-Enk in the nasal, rectal and vaginal mucosal extracts were 1.62, 0.37 and 1.12 hrs and those of YAGFL were 30.55, 9.70 and 6.82 hrs, respectively, indicating Leu-Enk was degraded in a more extensive and rapid manner than YAGFL. But the mucosal and serosal extracts of the same mucosa showed the similar degradation rates for both pentapeptides. The degradation was most rapid in the neutral pH and increasing concentrations of substrates retarded the degradation rates. The maior hydrolytic fragments of Leu-Enk were Des-Tyr-Leu-Enk and tyrosine, indicating the enzymatic hydrolysis by aminopeptidases. However, the data also suggested endopeptidases such as dipeptidyl carboxypeptidase and dipeptidyl aminopeptidase could play some role in the degradation of Leu-Enk. On the other hand, the hydrolytic fragments of YAGFL in all the mucosa extracts were mainly Tyr-D-Ala-Gly and Phe-Leu-Amide, demonstrating the hydrolytic breakdown by endopeptidases. The degradation pathways were further explored by concomitantly determining the formation of smaller metabolites of primary hydrolytic fragments of Leu-Enk and YAGFL in the mucosa extracts.

• Korean Journal of Weed Science
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• v.15 no.1
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• pp.73-84
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• 1995

Four malformin A's produced by Aspergillus niger van Tiegh. were separated by HPLC equipped with $C_{18}$ reversed-phase column and subjected to structural determination. Amino acid analyses and mass spectra data of the compounds indicate that they structurally resemble the cyclic pentapeptide malformin $A_1$. Their structures were deduced by two dimensional NMR and MS/MS experiments as cyclo-D-Cys-D-Cys-L-Val-D-Leu-L-Ile for $A_1$, cyclo-D-Cys-D-Cys-L-Val-D-Leu-L-Val for $A_2$, cyclo-D-Cys-D-Cys-L-Val-D-Leu-L-Leu for $A_3$, and cyclo-D-Cys-D-Cys-L-Val-D-Ile-L-Val for $A_4$. Among the mal-formin A's, the structure of $A_3$ was identical to that of malformin C, which was produced by A. niger strain AN-1. All the malformin A's caused severe curvatures of corn(Zea mays L.) roots and the activities of the malformin A's with molecular weight 529 were greater than those with molecular weight 515. Malformin $A_1$ caused the corn root curvature by 83% at a concentration of $0.25{\mu}M$. In the mung bean(Phaseolus aureus Roxb.) hypercotyl segment test, however, the molecular weight of malformin A's was not a factor influencing the physiological activities. Malformin $A_1$ stimulated the growth of mung bean hypercotyles by 165% at a $0.1{\mu}M$ concentration.

• Korean Journal of Weed Science
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• v.15 no.1
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• pp.85-98
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• 1995

Six malformin B's produced by Aspergillus niger van Tiegh. were separated by HPLC. Their structures determined by the methods of amino acid analyses, mass spectrometry, and two-dimensional NMR were revealed as cyclic pentapeptides structurally related to malformin $A_1$. Both the NMR and MS/MS data suggest that the respective structures of separated malformin B's were as follows; cyclo-D-Cys-D-Cys-L-Val-D-Leu-L-allo-Ile for $B_{1a}$, cyclo-D-Cys-D-Cys-L-Val-D-Leu-L-Leu for $B_{1b}$, cyclo-D-Cys-D-Cys-L-Val-D-Val-L-Leu for $B_2$, cyclo-D-Cys-D-Cys-L-Val-D-Ile-L-Leu for $B_3$, cyclo-D-Cys-D-Cys-L-Val-D-Ile-L-Ile for $B_4$, and cyclo-D-Cys-D-Cys-L-Val-D-Val-L-Ile for $B_5$. Among the malformin B's, the structure of $B_{1b}$ was the same as that of malformin $A_3$ or C. All the malformin B's showed physiological activities in the two assay systems using corn(Zea mays L.) roots and mung bean(Phaseolus aureus Roxb.) hypercotyl segments. The malformin B's with molecular weight 529 were more effective for inducing corn root curvature than those with molecular weight 515. The difference in molecular weight of malformin B's, i.e., the retention time on HPLC, results in the polarity change of the whole malformin molecule which affects the revealation of the malformin activities. In addition, the disulfide form of the malformin B's gives the rigidity of the molecule, whereas the combination of the fourth and the fifth amino acid residues provides the optimal three-dimensional configuration to the malformin receptor of plants. Presumably, these two factors are appeared to be essential for the greatest physiological activity of malformin B's. malformin $B_{1a}$ caused the corn root curvature by 90% at a concentration of $0.25{\mu}M$. However, such differential activities with molecular weight of 529 or 515 of malformin B's were not found in the mung bean hypercotyl segment test. Maximum stimulation of mung bean hypercotyl growth was observed at $0.1{\mu}M$ concentration of malformin B's. The growth of the segments treated with $B_5$ was 154% greater than that of the control.

• Fisheries and Aquatic Sciences
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• v.18 no.2
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• pp.131-135
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• 2015

Two new bitter-tasting cyclic peptides comprising six amino acids, namely gamakamide C and D, were isolated from cultured oysters Crassostrea gigas. Dimethylaminoazobenzene sulfonyl-amino acid analysis detected Val and Leu in gamakamide C and Ile and Leu in gamakamide D. The molecular formula of gamakamide C was determined as $C_{43}H_{60}N_{7}O_8S$ by high-resolution fast atom bombardment mass spectroscopy (HR FAB-MS) ($[M+H]^+m/z822.4200{\Delta}-2.4mmu$), and that of gamakamide D was determined as $C_{43}H_{62}N_7O_8S$ by HR FAB-MS ($[M+H]^+m/z836.4379{\Delta}-2.0mmu$). Comparison of amino acid analyses and fragment ions by MS/MS among gamakamide C, D, and E (known), the structures of gamakamide C and D were confirmed $as-{\small{L}}-Val-{\small{L}}-Met(SO)-{\small{L}}-NMe-Phe-{\small{L}}-Leu-{\small{D}}-Lys-{\small{L}}-Phe-$ and $-{\small{L}}-Ile-{\small{L}}-Met(SO)-{\small{L}}-NMe-Phe-{\small{L}}-Leu-{\small{D}}-Lys-{\small{L}}-Phe-$, respectively.

• Journal of Pharmaceutical Investigation
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• v.26 no.3
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• pp.175-185
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• 1996

To inhibit the enzymatic degradation of leucine enkephalin (Leu-Enk) and its synthetic analog. $[D-ala^2]$-leucine enkephalinamide (YAGFL), in the nasal, rectal and vaginal mucosal and serosal extracts of rabbits, effects of enzyme inhibitors such as amastatin (AM), puromycin (PM), thiorphan (TP), thimerosal (TM), EDTA, N-carboxymethyl-Phe-Leu (CPL), phenylethyl alcohol (PEA), phenylmercuric acetate (PMA), benzalkonium chloride (BC) and modified cyclodextrins, alone or in combination, were observed by assaying the pentapeptides staying intact during incubation. Mucosa extracts were prepared by exposing freshly-excised mucosal specimens mounted on Valia-Chien cells to isotonic phosphate buffer while stirring. The degradation of Leu-Enk and YAGFL followed the apparent first-order kinetics. The half-lives (mean) in the nasal, rectal and vaginal mucosal extracts were found to be 1.07, 0.33 and 1.14 hr for Leu-Enk, and 16.9, 6.2 and 6.8 hr for YAGFL, respectively. AM or PM, which is an aminopeptidase inhibitor, did not show a sufficient inhibition of Leu-Enk $(50\;{\mu}g/ml)$ degradation in all kinds of extracts. $Dimethyl-{\beta}-cyclodextrin\;(DM-{\beta}-CyD)$ decreased the degradation rate constants of Leu-Enk about 2 or 3 times, comparing with no additive. However, the use of mixed inhibitors of AM $(50\;{\mu}M)$/TM (0.25 mM)/EDTA (5 mM) resulted in a full stabilization of Leu-Enk by decreasing the degradation rate constants 67.3, 161.3 and 113.8 times far the nasal, rectal and vaginal mucosal extracts, respectively, comparing with no inhibitor. With mixed inhibitors, Leu-Enk remained intact more than 90% after 6 hr-incubation. In the stabilization of YAGFL, hM, TP or CPL alone showed little efffct, and some additives demonstrated a considerable inhibition of YAGFL degradation in the rank order of TM > BC > EDTA. However, the addition of mixed inhibitors such as TM (0.5 mM) and EDTA (5 mM) into the extracts protected YAGFL from the degradation by more than 85% even after 24 hr-incubation, suggesting almost complete inhibition of YAGFL degradation in the extract. On the other hand, $DM-{\beta}-CyD\;or\;hydroxypropyl-{\beta}-cyclodextrin$ (10%) were also found to retard enzymatic degradation rates of YAGFL markedly, and resulted in staying intact more than 80% of YAGFL in the nasal and vaginal mucosal extracts, and more than 60% in the rectal mucosal extract after 16 hr-incubation.

• YAKHAK HOEJI
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• v.38 no.5
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• pp.488-495
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• 1994

To evaluate the feasibility of transmucosal delivery of leucine enkephalin (Leu-Enk) and its synthetic analog, $[D-Ala^2]$-leucine enkephalinamide (YAGFL), their physicochemical stabilities in aqueous buffered solutions were first investigated using a stability indicating high performance liquid chromatography. The degradation of Leu-Enk and YAGFL followed the pseudo-first-order kinetics. From the pH-rate profiles, it was found that the maximal stability of the two pentapeptides was at the pH of about 5.0. The shelf lives $(t_{90%})$ for the degradation of Leu-Enk and YAGFL at pH 5.0 and $40^{\circ}C$ were found to be 48.13 and 50.9 days, respectively. From the temperature dependence of the degradation, activation energies for Leu-Enk and YAGFL were calculated to be 13.61 and 13.47 kcal/mole, respectively. A higher ionic strength and a higher initial peptide concentration in buffered solution slowed the degradation of the two pentapeptides. The addition of 2-hydroxypropyl-${\beta}$-cyclodextrin into the peptide solution did not affect the stability significantly.

• Microbiology and Biotechnology Letters
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• v.28 no.4
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• pp.195-201
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• 2000

Glucoamylase of Saccharomyces diastaticus is produced as a large precursor composed of signal peptide (21 amino acid residues), Thr and Ser-rich region and functional glucoamylase. To evaluate the utility of the glucoamylase signal peptide (GSP) for the secretion of foreign proteins, four types of GSP mutants (ml : Pro-18 longrightarrowLeu-18, m2 : Tyr-13 longrightarrowLeu, m3 : Ser-9longrightarrowLeu-9, m4 : Asn-5 longrightarrowPro-5) were constructed and secretion efficiency of each mutant was compared with that of native GSP by the expression and secretion of Bacillus subtilis CMCase under the control of GAP in N-terminal domain and hydrophobic domain. n mutant 4, a polar amino acid was replaced by a helix - breaking Pro residue. CMCase activity assay and Western blot analysis revealed that CMCase secretion by GSP mutants replaced by Leu were increased compared with native GSP. In the case of m2 and m3, the substitution of Leu for Tyr-13 and Ser-9 in the hydrophobic region resulted in a twofold increase in the extracellular CMCase activity.