• Title/Summary/Keyword: Serine Protease

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CHARACTERIZATION OF A HUMAN $\alpha_1$-ANTITRYPSIN VARIANT THAT IS AS STABLE AS OVALBUMIN BUT RETAINS INHIBITORY ACTIVITY

  • Lee, Kee-Nyung;Yu, Myeong-Hee
    • Proceedings of the Korean Biophysical Society Conference
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    • 1996.07a
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    • pp.14-14
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    • 1996

  • The metastable native state of proteins plays an important role in regulating biological functions. The native strain of serpins (serine protease inhibitors) are considered to be crucial for the inhibitory function. Several thermostable mutations of human $\alpha$$_1$-antitrypsin, a prototype inhibitory serpin, were identified in a systematic search targeted at the hydrophobic core of the molecule [Nature structural biology, vol. 3, no. 6, 497-500(1996)]. (omitted)

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Regulation of the Inhibitory Function of $\alpha_1$-Antitrypsin by Native Metastability

  • Lee, Cheolju;Yu, Myeong-Hee
    • Proceedings of the Korean Biophysical Society Conference
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    • 1999.06a
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    • pp.41-41
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    • 1999

  • The native forms of some proteins such as inhibitory serpins (serine protease inhibitors) and viral membrane fusion proteins are metastable, which is critical to their functions. To understand the mechanism of how native metastability regulates the inhibitory function of serpins, we characterized stabilizing mutations of $\alpha$$_1$-antitrypsin, a prototype serpin, in which Gly 117 was replaced by a series of larger hydrophobic residues.(omitted)

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Human $\alpha_1$-Antitrypsin Variant with Enhanced Conformational Stability at the Cost of Activity

  • Seo, Eun-Joo;Hana Im;Yu, Myeong-Hee
    • Proceedings of the Korean Biophysical Society Conference
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    • 1997.07a
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    • pp.39-39
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    • 1997

  • Native strain of inhibitory SERPINS (Serine protease inhibitors) is thought to be used in the facile conformational switch to play biological regulation. Many heat stable variants of $\alpha$$_1$-antitrypsin, a prototype of inhibitory serpins, increased their stability by reducing the native strain.(omitted)

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Fibrinolytic Activities and Effects of Gamma-Irradiated on Seeds from Coix lacryma-jobi L. Carthamus tinctorius L. and Malva verticillata L. (율무, 홍화, 아욱종자의 혈전용해 효소활성 및 감마선 조사의 영향)

  • Kwon Su-Jung;Lim Chae-Young;Kim Jae-Sung;Park Min-Hee;Lee Sook-Young
    • KSBB Journal
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    • v.21 no.1 s.96
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    • pp.20-27
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    • 2006

  • The fibrinolytic activities of soluble proteins extracted from seeds of Coix lacryma-jobi L., Carthamus tinctorius L. and Malva venicillata L. were studied. Fibrinolytic activity of extract from C. lacryma-jobi L. showed 1.3 times higher than plasmin used as positive control. The fibrinolytic enzyme was confirmed and extracted directly from seed of C. lacryma-jobi L. by a fibrin zymography. The protein was composed of a single polypeptide and its apparent molecular weight was found to be 7.8 kDa, as judged by SDS-polyacrylamide gel electrophoresis. The effect of temperature for the proteolytic enzyme activity were stabilized above $50^{\circ}C$ and then dramatically decreased. Also, the enzyme activity was clearly inhibited by APMSF, PMSF and TPCK, suggesting that it is a member of the chymotrypsin-like serine pretense. In addition, effects of gamma-irradiated on seed of each plants were revealed that 8 Gy and 64 Gy were higher than others. This result shown that gamma-irradiation of seeds were capable to increase the fibrinolytic activity. All these results suggest the pretense is a fibrinolytic enzyme belong to a family of chymotrypsin-like serine pretense.

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Purification and Characterization of a Novel Alkaline Protease from Bacillus horikoshii

  • Joo, Han-Seung;Choi, Jang-Won
    • Journal of Microbiology and Biotechnology
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    • v.22 no.1
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    • pp.58-68
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    • 2012

  • An investigation was conducted on the enhancement of production and purification of an oxidant and SDS-stable alkaline protease (BHAP) secreted by an alkalophilic Bacillus horikoshii, which was screened from the body fluid of a unique Korean polychaeta (Periserrula leucophryna) living in the tidal mud flats of Kwangwha Island in the Korean West Sea. A prominent effect on BHAP production was obtained by adding 2% maltose, 1% sodium citrate, 0.8% NaCl, and 0.6% sodium carbonate to the culturing medium. The optimal medium for BHAP production contained (g/l) SBM, 15; casein, 10; $K_2HPO_4$, 2; $KH_2PO_4$, 2; maltose, 20; sodium citrate, 10; $MgSO_4$, 0.06; NaCl, 8; and $Na_2CO_3$, 6. A protease yield of approximately 56,000 U/ml was achieved using the optimized medium, which is an increase of approximately 5.5-fold compared with the previous optimization (10,050 U/ml). The BHAP was homogenously purified 34-fold with an overall recovery of 34% and a specific activity of 223,090 U/mg protein using adsorption with Diaion HPA75, hydrophobic interaction chromatography (HIC) on Phenyl-Sepharose, and ion-exchange chromatography on a DEAE- and CM-Sepharose column. The purified BHAP was determined a homogeneous by SDS-PAGE, with an apparent molecular mass of 28 kDa, and it showed extreme stability towards organic solvents, SDS, and oxidizing agents. The $K_m$ and $k_{cat}$ values were 78.7 ${\mu}M$ and $217.4s^{-1}$ for N-succinyl-Ala-Ala-Pro-Phe-pNA at $37^{\circ}C$ and pH 9, respectively. The inhibition profile exhibited by PMSF suggested that the protease from B. horikoshii belongs to the family of serine proteases. The BHAP, which showed high stability against SDS and $H_2O_2$, has significance for industrial application, such as additives in detergent and feed industries.

  • https://doi.org/10.4014/jmb.1109.09006 인용 PDF KSCI