• 셀메드
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• 제4권1호
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• pp.6.1-6.12
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• 2014

Anthrax is the deadly disease for human being caused by Bacillus anthracis. Instantaneous research work on the mode of infection of the organism revealed that different proteases are involved in different steps of pathogenesis. Present study reports the in silico characterization and the detection of pathogenic proteases involved in anthrax infection through protein-protein interaction. A total of 13 acid, 9 neutral, and 1 alkaline protease of Bacillus anthracis were selected for analysing the physicochemical parameter, the protein superfamily and family search, multiple sequence alignment, phylogenetic tree construction, protein-protein interactions and motif finding. Among the 13 acid proteases, 10 were found as extracellular enzymes that interact with immune inhibitor A (InhA) and help the organism to cross the blood brain barrier during the process of infection. Multiple sequence alignment of above acid proteases revealed the position 368, 489, and 498-contained 100% conserved amino acids which could be used to deactivate the protease. Among the groups analyzed, only acid protease were found to interact with InhA, which indicated that metalloproteases of acid protease group have the capability to develop pathogenesis during B. anthracis infection. Deactivation of conserved amino acid position of germination protease can stop the sporulation and germination of B anthracis cell. The detailed interaction study of neutral and alkaline proteases could also be helpful to design the interaction network for the better understanding of anthrax disease.

• 한국식품과학회지
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• 제46권6호
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• pp.702-709
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• 2014

Protease 무첨가가 숙성약주의 이화학적 특성에 미치는 영향을 조사하였다. 발효온도 $20-25^{\circ}C$에서는 정상적으로 알코올 생산을 하였으나 발효온도 $10^{\circ}C$에서는 첨가구보다 21.3%, $15^{\circ}C$에서는 9.4%, $30^{\circ}C$에서는 13.2% 낮은 알코올 생산성을 보여주었다. 발효기간은 발효온도 $25^{\circ}C$를 제외하고 protease 첨가구 대비 증가하는 것으로 나타났다. 총 유기산 함량은 1.0-2.7배 증가하였으며, lactic acid와 acetic acid가 증가된 반면 pyroglutamic acid의 함량이 최대 51.1배 감소되었다. 산도와 휘발산은 증가되었고 pH는 감소되는 것으로 나타났다. 아미노산도는 protease 첨가구에 비하여 3.0-5.2배, 유리 아미노산 총량은 5.2-11.9배 낮게 나타났다. Protease 첨가구의 주요 유리 아미노산은 alanine, arginine, leucine, lysine인 반면 무첨가구는 alanine, arginine, leucine, phenylalanine 이었다. 색차는 protease 첨가구에 비해 1.2-3.0배 낮아졌다.

• 한국식품영양과학회지
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• 제13권2호
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• pp.193-204
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• 1984

Zymolyase 조효소로부터 분리, 정제되었고, 효모세포벽 용해 촉진물질로 밝혀진 바 있는 Arthrobacter luteus로부터 유래한 염기성 protease(AL-protease)의 효소학적 성질 및 활성 아미노산 잔기를 검색한 결과는 다음과 같다. 1. AL-pretense는 저해제 DFP 및 PMSF에 의해서 그 Protease 활성 및 용해 촉진활성이 동시에 완전히 소멸 되었으며, 그 저해 반응속도는 chymotrypsin에 대한 것에 비하여 대단히 완만하였다. 1.반응에서 AL-protease와 DFP의 결합 mole비는 1:1로 추정 되었다. 2. 정제된 AL-protease의 동결건조품 중에는 종래효모세포벽 용해반응에 관여하는 것으로 알려진 yeast phosphomannase를 비롯한 다당류 가수분해효소들의 활성은 그 어느 것도 인정되지 않았다. 3. AL-protease의 casein에 대한 최적 pH 및 최적 온도는 pH 10.5와 $65^{\circ}C$이었고, 그 활성은 pH 5${\sim}$11 사이와 $65^{\circ}C$이하에서 안정하였다. 또한, AL-Protease의 활성에 미치는 여러가지 금속이온의 영향은 인정되지 않았다. 4. [$^{32}P$]-DFP에 의하여 화학수식된 [$^{32}P$]-DIP-AL-protease에 대한 활성부위의 아미노산 잔기를 검색, 동정하기 위하여 조제용 PAG-전기영동, SDS-PAG-전기영동, Dowex 이온교환 크로마토그래피 및 고압 여지 전기영동을 실시하였고, 그 결과, AL-protease는 활성 부위에 1분자당 1 mole의 serine 잔기를 가지는 염기성 protease로 밝혀졌다.

• International journal of advanced smart convergence
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• 제10권1호
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• pp.197-208
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• 2021

The deer antler has been used as a major drug in oriental medicine for a long time. Recently, the demand for easy-to-take health functional foods is increasing due to economic development and changes in diet. As part of research on the development of functional materials for antlers, lactic acid fermentation of antler extract was performed. It was intended to develop a functional material with enhanced total polyphenol and flavonoid content and enhanced antioxidant activity. Lactic acid bacteria fermentation was performed by adding 4 types of lactic acid bacteria starter products, B. longum, Lb. Plantarum, Lb. acidophilus and mixture of 8 types of lactic acid bacteria to the antler water extract substrate, respectively. During the fermentation of lactic acid bacteria, the number of proliferation, total polyphenol and total flavonoid content, DPPH radical scavenging and antioxidant activity were quantified and evaluated. As a result of adding these four types of lactic acid bacteria to the antler water extract substrate, the number of lactic acid bacteria measured was 2.04~5.00×107. Meanwhile, a protease (Baciullus amyloliquefaciens culture: Maxazyme NNP DS) was added to the antler extract to decompose the peptide bonds of the contained proteins. Then, these four types of lactic acid bacteria were added and the number of lactic acid bacteria increased to 2.84×107 ~ 2.21×108 as the result of culture. The total polyphenol contents were 4.82~6.26 ㎍/mL in the lactic acid bacteria fermentation extracts, and after the reaction of protease enzyme and lactic fermentation, increased to 14.27~20.58 ㎍/mL. The total flavonoid contents were 1.52~2.21 ㎍/ml in the lactic acid bacteria fermentation extracts, and after the protease reaction and fermentation, increased to 5.59 ~ 8.11 mg/mL. DPPH radical scavenging activities of lactic acid bacteria fermentation extracts was 17.03~22.75%, but after the protease reaction and fermentation, remarkably increased to 32.82~42.90%.

• BMB Reports
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• 제37권5호
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• pp.574-581
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• 2004

The full-length cDNA of the lumbrokinase fraction 6 (F6) protease gene of Lumbricus rubellus was amplified using an mRNA template, sequenced and expressed in E. coli cells. The F6 protease gene consisted of pro- and mature sequences by gene sequence analysis, and the protease was translated and modified into active mature polypeptide by N-terminal amino acid sequence analysis of the F6 protease. The pro-region of F6 protease consisted of the 44 residues from methionine-1 to lysine-44, and the mature polypeptide sequence (239 amino acid residues and one stop codon; 720 bp) started from isoleucine-45 and continued to the terminal residue. F6 protease gene clones having pro-mature sequence and mature sequence produced inclusion bodies in E. coli cells. When inclusion bodies were orally administrated rats, generated thrombus weight in the rat' venous was reduced by approximately 60% versus controls. When the inclusion bodies were solubilized in pepsin and/or trypsin solutions, the solubilized enzymes showed hemolytic activity in vitro. It was concluded the F6 protease has hemolytic activity, and that it is composed of pro- and mature regions.

• 한국동물학회지
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• 제27권4호
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• pp.199-212
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• 1984

대장균의 세포질내에 존재하는 단백질 분해효소(protease Ci)를 추출하여 그 성질을 조사하였다. 이 효소는 insulin, glucagon 및 bovine growth hormone을 분해시키나, bovine serum albumin, casein 및 globin 등은 분해시키지 않는다. 이 효소의 native 한 상태에서의 분자량은 120,000이며, 54,000 dalton의 동일한 2개의 subunit로 구성되어 있다. Protease Ci의 최적 pH는 7.5이며 등전점은 5.5이다. 이 효소는 o-phenanthroline에 의해 저해되며, $Mn^++나 Co^++$와 같은 metal 이온들에 의하여 활성화되므로 metalloprotease임을 알 수 있다. 이 효소는 p-hydroxymercuribenzoate에 의해서 크게 저해되나 sulfhydryl protease의 specific한 저해제인 Ep475와 leupeptin에 의해서는 영향을 받지 않는다. 대장균 세포내에서는 또 다른 insulin 분해효소 (protease Pi)가 존재하는데, 이 효소는 periplasm에 존재하므로 protease Ci와는 다르다.

• 미생물학회지
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• 제18권2호
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• pp.51-58
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• 1980

Mutational experiments were performed to improved to improve the protease productivity of Aspergillus flavus KU 153, which is selected among the wild strains. A UV-induced mutant strain having high protease productivity was obtained by the use of the clear zone method as a simple criterion for a primary screening test. Neutral and alkaline protease activities of hte mutant strain were higher than 1.8 times, comopared with those of the parental strain, respectively, while in the case of acid protease, it was 2.7 times. The mutant strain selected was more powerful in the production of cellulase and amylase, as well s protease in wheat bran, compared with those of the parental strain. protease production of the parental strain has reached maximum level at 3 days culture, while alkaline nad neutral protease production of the mutantstrain has reached at 2 days culture. On the other hand, the mutant strain formed the spore slowly, compared with the parental strain. Column chromatography of the neutral protease on DEAE-Sephadex A-50 showed that the mutant strain was not induced the formation of another neutral protease isozyme, but induced the variation in the function of regulatory gene.

• 한국식품영양학회지
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• 제19권4호
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• pp.496-501
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• 2006

본 연구는 대한민국 서해안 갯벌로부터 알칼리성 단백질 분해 효소를 생산하는 세균을 분리하고 분리된 세균으로부터 생산되는 단백질 분해 효소의 생화학적 특징을 조사하는 것이다. 분리 균주는 16S rRNA의 염기 서열, 그람 염색과 전자 현미경사진을 통해 Celidibacter sp. HK-1으로 명명하였다. 분리 균주의 증식과 pretense 생산을 위한 최적 온도는 $25^{\circ}C$이었다. 분리 균주의 증식은 접종 10시간후에 stationary phase에 도달하였다. 효소 생산은 14시간 후 최대 값을 보였다. 효소 활성의 최적 온도와 pH는 각각 $45^{\circ}C$와 pH 9이었다. Pretense의 분자량은 약 50KD이었고 pretense의 부분적인 아미노산 서열은 Ala-Try-Ala-Leu-Asn-Thr-Ser-Val-Thr-Glu-Thr-Phe-Ala-Lys이었다. Protease의 부분적인 아미노산 서열은 Streptomyces avemitilis의 pretense와 높은 상동성을 보였다.

• Journal of Microbiology
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• 제33권2호
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• pp.103-108
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• 1995

Streptomyces fradiae protease inhibitor (SFI) was purified effectively by preparative isoelectric focusing and hydroxyapatite chromatography. The molecular weight of SFI was estimated to be 1.7 kDa by SDS-PAGE and 1.8 kDa by molecular sieving HPLC. One hundred and sixty amino acid residues were determined from which molecular weight of SFI was calculated to be 17.054 Da and carbohydrate residue was not detected. SFI was calculated to be 17,064 Da and carbohydrate residue was not detected. SFI was a monomeric protein with two reactive sits, of which isoelectric point was pH 4.1. N-terminal amino acid sequence of SFI had homology with SSI (Streptomyces subsilisin inhibitor) and other protease inhibitors produced by Streptomyces.

• Applied Biological Chemistry
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• 제33권4호
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• pp.325-329
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• 1990

젓갈에서 분리한 B. subtilis p-4와 B. licheniformis p-5 프로테아제의 기질(카제인)에 대한 친화도를 측정한 결과 각각 0.38mM, 0.18mM이었고 열변성에 의한 특성을 반응속도론적으로 검토한 결과, p-4 프로테아제는 $50^{\circ}C$에서 80분 열처리할 때 20%의 잔존 활성을 보였고, p-5 프로테아제는 거의 실활되었는데 일차반응식을 따랐다. 그리고 p4, p-5 프로테아제의 변성 속도 상수는 각각 $40^{\circ}C$에서 $12.2{\times}10^{-5}/sec,\;19.0{\times}10^{-5}/sec,\;50^{\circ}C$에서는 $35.7{\times}10^{-5}/sec,\;46.3{\times}10^{-5}/sec$였다. 또 활성화 에너지는 p-4프로테아제가 19.6Kca1/mole, p-5 프로테아제는 15.2Kcal/mole로 p-5 프로테아제가 열변성에 민감한 것을 알 수 있었다. 활성화 자유에너지는 p-4, p-5 프로테아제 모두 온도가 상승함에 따라 약간 증가하였는데 $40^{\circ}C$에서 각각 23.21, 22.93Kcal/mole, $50^{\circ}C$에서는 23.28, 23.11Kcal/mole이었다. 아미노산 조성은 p-4 프로테아제의 경우 151개의 아미노산 잔기를 가졌으며, 이중 glycine, glutamic acid, proline이 많았고, p-5 프로테아제는 247개의 잔기에 glycine, glutamic acid, aspartic acid가 많았다.