Bacillus sp. EBTA6 ile Proteaz Üretimi için Plackett–Burman Dizaynı Kullanılarak Besiyeri Bileşenlerinin Taranması ve Seçimi

Bu çalışmada, Plackett–Burman Dizaynı kullanılarak ev yapımı bir Tarhanadan izole edilen Bacillus sp. EBTA6’nın proteaz üretimine besiyeri bileşenleri ve inokülüm miktarının etkisi araştırılmıştır. Yanıt olarak proteaz aktivitesinin belirlenmesi amacıyla çalkalamalı inkübatörde 24 saat geliştirilen bakterinin hücresiz süpernatantı kullanılmıştır. Toplam 11 faktör ile 12 farklı deneme gerçekleştirilmiş ve en yüksek deneysel proteaz aktivitesi 2280,4 U/mL olarak ölçülmüştür. Sonuçlar ANOVA ile istatistiki olarak analiz edilmiş ve en etkili faktörler %93,78; 2,19; 1,96; 1,31 katkı oranları ile sırasıyla maya özütü, dipotasyum fosfat, kazein ve pepton olarak tespit edilmiştir. Seçilen faktörlerin doğrulanması için dizayn denkleminden elde edilen maya özütü (9,98 g/L), dipotasyum fosfat (1,27 g/L), kazein (8,69 g/L) ve pepton (9,88 g/L) ile de bir deneme gerçekleştirilmiştir. Deneysel yanıt, tahmin edilen yanıttan yalnızc

Screening and Selection of Media Components for Protease Production by Bacillus sp. EBTA6 Using Plackett–Burman Design

In this study, effects of medium components and inoculum size on the protease production by Bacillussp. EBTA6 that was isolated from a home-made Tarhana sample were investigated. The cell-freesupernatant of bacterium cultured on a shaking incubator for 24 h was used to test protease activityas the response. With a total number of 11 factors, 12 different experiments were run and the highestexperimental protease activity was measured as 2280.4 U/mL. Results were analyzed statistically byANOVA and the most efficient factors were detected as yeast extract, dipotassium phosphate, casein,and peptone with a contribution of 93.78, 2.19, 1.96, 1.31%, respectively. For validation of theselected factors, a further experiment was performed by using of yeast extract (9.98 g/L), dipotassiumphosphate (1.27 g/L), casein (8.69 g/L), and peptone (9.88 g/L) obtained from the design equation.The experimental response was found as 2411.4 U/mL which was only 5.5% higher than the predictedresponse showing that the model was applicable.

PDF

___

Banik RM, Prakash M. 2004. Laundry detergent compatibility of the alkaline protease from Bacillus cereus. Microbiol. Res., 159: 135-140. DOI: 10.1016/j.micres.2004.01.002
Beg QK, Sahai V, Gupta R. 2003. Statistical media optimization and alkaline protease production from Bacillus mojavensis in a bioreactor. Process Biochem., 39: 203-209. DOI: 10.1016/S0032-9592(03)00064-5
Białkowska AM, Krysiak J, Florczak T, Szulczewska KM, Wanarska M, Turkiewicz, M. 2018. The psychrotrophic yeast Sporobolomyces roseus LOCK 1119 as a source of a highly active aspartic protease for the in vitro production of antioxidant peptides. Biotechnol. Appl. Biochem., 65: 726- 738. DOI: 10.1002/bab.1656
Borhani MS, Etemadifar Z, Emtiazi G, Jorjani E. 2018. A statistical approach for production improvement of a neutral protease from a newly isolated strain of Aeromonas hydrophil. Iran. J. Sci. Technol. Trans. Sci., 42: 1771–1778. DOI: 10.1007/s40995-017-0444-1
Chauhan B, Gupta R. 2004. Application of statistical experimental design for optimization of alkaline protease production from Bacillus sp. RGR-14. Process Biochem., 39: 2115-2122. DOI: 10.1016/j.procbio.2003.11.002
Ferrero MA, Castro GR, Abate CM, Baigori MD, Sineriz F. 1996. Thermostable alkaline proteases of Bacillus licheniformis MIR 29: isolation, production and characterization. Appl. Microbiol. Biotechnol., 45: 327-332. DOI: 10.1007/s002530050691
Ghosh P, Mukherji S. 2018. Optimization of media composition for enhancing carbazole degradation by Pseudomonas aeruginosa RS1. J. Environ. Chem. Eng., 6: 2881-2891. DOI: 10.1016/j.jece.2018.04.043
Gupta R, Beg Q, Lorenz P. 2002. Bacterial alkaline proteases: molecular approaches and industrial applications. Appl. Microbiol. Biotechnol., 59: 15-32. DOI: 10.1007/s00253- 002-0975-y
Hammami A, Bayoudh A, Abdelhedi O, Nasri M. 2018. Low-cost culture medium for the production of proteases by Bacillus mojavensis SA and their potential use for the preparation of antioxidant protein hydrolysate from meat sausage byproducts. Ann. Microbiol., 68: 473-484. DOI: 10.1007/s13213-018-1352-0
Horikoshi, K. (1999). Alkaliphiles: some applications of their products for biotechnology. Microbiol. Mol. Biol. Rev., 63: 735-750.
Ibrahim ASS, Al-Salamah AA, Elbadawi YB, El-Tayeb MA, Ibrahim SSS. 2015. Production of extracellular alkaline protease by new halotolerant alkaliphilic Bacillus sp. NPSTAK15 isolated from hyper saline soda lakes. Electron. J. Biotechnol., 18: 236-243. Available from: https://scielo.conicyt.cl/scielo.php?pid=S0717- 34582015000300015&script=sci_arttext [20.09.2019]. DOI: 10.1016/j.ejbt.2015.04.001
Joo HS, Kumar CG, Park GC, Kim KT, Paik SR, Chang CS. 2002. Optimization of the production of an extracellular alkaline protease from Bacillus horikoshii. Process Biochem., 38: 155-159. DOI: 10.1016/S0032-9592(02)00061-4
Karlapudi AP, Krupanidhi S, Reddy ER, Indira M, Bobby MdN, Venkateswarulu TC. 2018. Plackett-Burman design for screening of process components and their effects on production of lactase by newly isolated Bacillus sp. VUVD101 strain from Dairy effluent. Beni-Seuf Univ. J. Appl. Sci., 7: 543-546. DOI: 10.1016/j.bjbas.2018.06.006
Kaur K, Sondhi S, Kaur PS. 2017. Statistical optimization of the production of protease from Bacillus sp. MSK-01 in submerged fermentation. J. Commer. Biotechnol., 23: 28-37. DOI:10.5912/jcb810
Kembhavi AA, Kulkarni A, Pant A. 1993. Salt-tolerant and thermostable alkaline protease from Bacillus subtilis NCIM No. 64. Appl. Biochem. Biotechol., 38: 83-92. DOI: 10.1007/BF02916414
Lim J, Choi YH, Hurh BS, Lee I. 2019. Strain improvement of Aspergillus sojae for increased L-leucine aminopeptidase and protease production. Food Sci. Biotechnol., 28: 121-128. DOI: 10.1007/s10068-018-0427-9
Limkar MB, Pawar SV, Rathod VK. 2019. Statistical optimization of xylanase and alkaline protease co-production by Bacillus spp. using Box-Behnken Design under submerged fermentation using wheat bran as a substrate. Biocatal. Agric. Biotechnol., 17: 455-464. DOI: 10.1016/j.bcab.2018.12.008
Moon SH, Parulekar SJ. 1991. A parametric study ot protease production in batch and fed‐batch cultures of Bacillus firmus. Biotechnol. Bioeng., 37: 467-483. DOI: 10.1002/bit.260370509
Patil U, Mokashe N, Shaha J, Arthekar S, Jagatap H. 2018. Ultrasound-assisted improvements in biocatalytic activity and production of organic-solvent stable protease from Bacillus circulans MTCC 7942. Ultrason. Sonochem., 40: 201-205. DOI: 10.1016/j.ultsonch.2017.07.012
Puri S, Beg QK, Gupta R. 2002. Optimization of alkaline protease production from Bacillus sp. by response surface methodology. Curr. Microbiol., 44: 286-290. DOI: 10.1007/s00284-001-0006-8
Rao MB, Tanksale AM, Ghatge MS, Deshpande VV, 1998. Molecular and biotechnological aspects of microbial proteases. Microbiol. Mol. Biol. Rev., 62: 597-635.
Rao RR, Vimudha M, Kamini NR, Gowthaman MK, Chandrasekran B, Saravanan P. 2017. Alkaline protease production from Brevibacterium luteolum (MTCC 5982) under solid-state fermentation and its application for sulfidefree unhairing of cowhides. Appl. Biochem. Biotechol., 182: 511-528. DOI: 10.1007/s12010-016-2341-z
Rekik H, Jaouadi NZ, Gargouri F, Bejar W, Frikha F, Jmal N, Bejar S, Jaouadi B. 2019. Production, purification and biochemical characterization of a novel detergent-stable serine alkaline protease from Bacillus safensis strain RH12. Int. J. Biol. Macromol., 121: 1227-1239. DOI: 10.1016/j.ijbiomac.2018.10.139
Shabbiri K, Adnan A, Jamil S, Ahmad W, Noor B, Rafique HM. 2012. Medium optimization of protease production by Brevibacterium linens DSM 20158, using statistical approach. Braz. J. Microbiol., 43: 1051-1061. DOI: 10.1590/S1517-83822012000300031
Sharma KM, Kumar R, Panwar S, Kumar A. 2017. Microbial alkaline proteases: Optimization of production parameters and their properties. J. Genet. Eng. Biotechnol., 15: 115-126. DOI: 10.1016/j.jgeb.2017.02.001
Si JB, Jang EJ, Charalampopoulos D, Wee YJ. 2018. Purification and characterization of microbial protease produced extracellularly from Bacillus subtilis FBL-1. Biotechnol. Bioproc. E., 23: 176-182. DOI: 10.1007/s12257-017-0495-3
Sudar M, Valinger D, Findrik Z, Vasić-Rački Đ, Kurtanjek Ž. 2013. Effect of different variables on the efficiency of the Baker's yeast cell disruption process to obtain alcohol dehydrogenase activity. Appl. Biochem. Biotechol., 169: 1039-1055. DOI: 10.1007/s12010-012-0056-3
Tari C, Genckal H, Tokatlı F. 2006. Optimization of a growth medium using a statistical approach for the production of an alkaline protease from a newly isolated Bacillus sp. L21. Process Biochem., 41: 659-665. DOI: 10.1016/j.procbio. 2005.08.012