Mikrobiyal Dipeptidil Peptidaz IV’ün Kinetik Özelliklerinin Belirlenmesi

Dipeptidil peptidaz IV (EC 3.4.14.5; DPPIV) enzimi tüm doku, organ ve sistemlerin düzenlenmesinde rol alan, nörolojik, immünolojik ve gastroenterolojik açıdan araştırılan önemli bir enzimdir. DPPIV enzimi pek çok memeli, bitki, böcek ve mikrobiyal hücrelerden regüle edici enzim olarak elde edilmektedir. DPPIV enziminin sindirim enzimi eksikliği, otizm, çölyak ve Asperger sendromu gibi hastalıklarda gıda takviyesi olarak kullanımıyla son yıllarda ticari olarak üretilmesi artmıştır. DPPIV enzim kaynağı olarak hücre-dışı enzim ekspresyonu yapan fungal kaynaklar tercih edilmesine rağmen bakterilerin enzim üretim verimliliği funguslara göre daha fazla olduğundan son yıllardaki ticari enzim üretimlerinde bakteriler tercih edilmektedir. Bu çalışmada, koleksiyon kültüründen satın alınmış bir laktik asit bakterisi, Lactococcus lactis spp. lactis (L. lactis), enzim üreticisi mikroorganizma olarak seçilmiştir. L. lactis, kazein proteinini hidrolizlemek için endo- ve ekzo-peptidazlar eksprese eder. DPPIV enzimi de bunlardan biridir. Hücre-içi DPPIV enzimi, üretiminden sonra saflaştırılmış ve ardından enzim karakterizasyonunu yapmak için kinetik özellikleri incelenmiştir. Sonuçlar, Gly-Pro-pNA substratı için optimum sıcaklığın 40 °C ve optimum pH’ nın 8,0 olduğunu göstermektedir. Saflaştırılan enzimin Gly-Pro-pNA substratına ilgisi olduğu bulunmuştur. Enzim, Hg+2, Zn+2, Ca+2, Fe+2 ve Mg+2 iyonları tarafından sırası ile inhibe edilmiştir. Enzim, Diprotin A tarafından güçlü bir şekilde inhibe edilmiştir. Jelatin inhibisyonu fenilmetilsulfonil florid (PMSF) inhibisyonu ile yakın bulunmuş ve etilendiamin tetra asetik asitin (EDTA) inhibisyona etkisi gözlenmemiştir.

___

  • Banbula A, Bugno M, Goldstein J, Yen J, Nelson D, Travis J, Potempa J (2000). Emerging family of proline-specific peptidases of Porphyromonas gingivalis: purification and characterization of serine dipeptidyl peptidase, a structural and functional homologue of mammalian prolyl dipeptidyl peptidase IV. Infection and immunity, 68(3): 1176-1182.
  • Baykal Y, Akyol T, ‘Romatoid Artritte Yeni Tedavi Yaklaşımları’ http://www.gata.edu.tr/dahilibilimler /ichastaliklari/files/kitaplar/137.pdf (Erişim Tarihi 29 Mayıs 2011)
  • Brandt I, Lambeir AM, Maes MB, Scharpé S, Meester ID (2006). Peptidesubstrates of dipeptidyl peptidases, dipeptidyl aminopeptidases. basic science andclinical applications, Advances in Experimental Medicine and Biology, 575 (18: 3-18.
  • Brudnak M, Buchholz I, Hoener S, Newman L, Pangborn J (2005). Guide to intestinal health in autism spectrum disorder, a comprehensive review of intestinal health issues in autism spectrum disorders and the options available for treating them, authored by Kirkman labs‟ Technical Staff, 1–175.
  • Buckley SJ, Collins PJ, O’Connor BF (2004). The purification and characterisation of novel dipeptidyl peptidase IV-like activity from bovine serum. The International Journal of Biochemistry and Cell Biology, 36(7): 1281-1296.
  • Çetin EM, Üstün Ö, Kırılmaz L, Öngen G (2007). The application of hydroxypropylmethylcellulose phtalate capsules: an alternative administration route of dipeptidyl peptidase enzyme. Pharmaceutical Sciences World Congress,22-25 April 2007, 63.Amsterdam.
  • Degraeve P and Martial-Gros A (2003). Purification and partial characterisation of xprolyl dipeptidyl aminopeptidase of Lactobacillus helveticus ITG LH1. International journal of Dairy Technology, 13(7): 497-507.
  • Elgün S, Keskinege A, Kumbasar H, (1999). Dipeptidyl peptidase IV and adenosine deaminase activity, decrease in depression, Psychoneuroendocrinology, 24:823-832.
  • Goldstein JM, Banbula A, Kordula T, Mayo JA, Travis J (2001). Novel extracellular x-prolyl dipeptidyl-peptidase (DPP) from Streptococcus gordonii FSS2: an emerging subfamily of viridans streptococcal x-prolyl dpps, Infection and Immunity, 69(9): 5494-5501.
  • Iwata S and Morimotoa CB (2007). CD26/Dipeptidyl Peptidase IV in Context, The Different Roles Of A Multifunctional Ectoenzyme in Malignant Transformation, Division Of Tumor Immunology, Dana-Farber Cancer Institute and Department of Medicine, Harvard Medical School, Boston, Ma 02115.
  • Janecki DJ and Reilly JP (2005). Denaturation of metalloproteins with EDTA to facilitate enzymatic digestion and mass fingerprinting, Rapid Communications in Mass Spectrometry, 19(10): 1268-1272.
  • Jobin MC,and Grenier D (2003). Identification and characterization of four proteases produced by Streptococcus suis. FEMS Microbiology Letters, 220(1): 113-119.
  • Jobin MC, Martinez G, Motard J, Gottschalk M, Grenier D (2005). Cloning, purification, and enzymatic properties of dipeptidyl peptidase IV from the swine pathogen Streptococcus suis. Journal of Bacteriology, 187(2): 795-799.
  • Kınık Ö ve Gürsoy O (2002). Süt proteinleri kaynaklı biyoaktif peptitler. Pamukkale Üniversitesi Mühendislik Fakültesi Mühendislik Bilimleri Dergisi, 8(2): 195- 203.
  • Maes M, Meester I, Scharpe S, Desnder R, Ranjan R, Meltzer Hy (1996). Alterations in plasma dipeptidyl peptidase ıv enzyme activity in depression and schizophrenia: effects of antidepressants and antipsychotic drugs, Acta Psychiatrica Scandinavica, 93:1-8.
  • Marchetti B, Scifo R, Batticane N, Scapagnini U (1990). Immunological significance of opioid peptide dysfunction in infantile autism, Brain Dysfunction, 3: 346-354.
  • Noni I, FitzGerald RJ, Korhonen HJT, Roux Y, Livesey CT, Thorsdottir I, Tomé D, Witkamp R (2009). Review of The Potential Health Impact of β-Casomorphins and Related Peptides. EFSA Scientific Report, 231: 1-107.
  • Perez Guzman EA, Victoria TC, Cruz-Camarillo R, Hernandez-Sanchez H (2004). Improvement of fermentetion conditions for the production of x-prolyl-dipeptidyl aminopeptidase from Lactococcus lactis, World Journal of Microbiology and Biotechnology, 20: 413-417.
  • Perez Guzman EA, Victoria TC, Cruz-Camarillo R, Hernandez-Sanchez H (2006). Purification and characterization of x-prolyl-dipeptidyl aminopeptidase from Lactococcus lactis subsp. cremoris NRRL 634, World Journal of Microbiology and Biotechnology, 22(9): 953-958.
  • Pihlanto-Leppälä A (2001). Bioactive pepdites derived from bovine whey proteins: opioid and ace-inhibitory pepdites, Trends in Food Scicence and Technology, 11: 347-356.
  • Saatci E, Erarslan A, Dinç H, İşcan A (2011). Sütteki şeytan: hastalıkta ve sağlıkta A1 ve A2 süt. 23. Ulusal Biyokimya Kongresi, 29 Kasım-2 Aralık 2011, s.36. Adana.
  • Sanz Y and Toldra F (2001). Purification and characterization of an x-prolyl-dipeptidyl peptidase from Lactobacillus sakei. Applied and Environmental Microbiology,, 67;1815-1820.
  • Sentandreu MA, Toldra F (2001). “Dipeptidyl Peptidase IV from Porcine Skeletal Muscle: Purification and Biochemical Properties”, Food Chem, 75, 2, 159–168.
  • Sigma (2006) In: Quality control procedure, quality control procedure enzymatic assay of dipeptidyl peptidase IV (EC 3.4.14.5), pp 1–2, Germany.
  • Simova E, Beshkova D, Angelov A, Hristozova T, Frengova G, Spazov Z (2002). Lactic acid bacteria and yeasts in kefir grains and kefir made from them, Journal of Industrial Microbiology and Biotechnology, 28 :1-6.
  • Sromova L, Mareckova H, Sedova L, Balaziova E, Sedo A (2010). Dipeptidyl peptidase-IV in synovial fluid and in synovial fluid mononuclear cells of patients with rheumatoid arthritis, Clinica Chimica Acta, 111:1046- 1050.
  • Stressler T, Eisele T, Schlayer M, Lutz-Wahl S, Fischer L (2013). Characterization of the recombinant exopeptidases PepX and PepN from Lactobacillus helveticus ATCC 12046 important for food protein hydrolysis, PloS one, 8(7): e70055.
  • Üstün-Aytekin Ö, Arısoy S, Aytekin AÖ, Yıldız E (2016). Statistical optimization of cell disruption techniques for releasing intracellular X-prolyl dipeptidyl aminopeptidase from Lactococcus lactis spp. Lactis, Ultrasonics Sonochemistry, 29: 163-171.
  • Üstün Ö ve Öngen G (2012). Production and separation of dipeptidyl peptidase IV from Lactococcus lactis: Scale up for industrial production, Bioprocess and Biosystems Engineering, 35 (8):1417-1427.
  • Van WD, Monteleone P, Di LA, Meester I, Durinx C, Scharpe S (2000). Lowered serum dipeptidyl peptidase ıv activity in patients with anorexia and blumia nervosa, European Archives of Psychiatry and Clinical Neuroscience, 250: 86-92.
  • Yan TR, Ho SC, Hou CL (1992). Catalytic properties of X-prolyl dipeptidyl aminopeptidase from Lactococcus lactis subsp. cremoris nTR, Bioscience, Biotechnology, and Biochemistry, 56(5): 704-707.
  • Zevaco C, Monnet V, Gripon JC (1990). Intracellular X-prolyl dipeptidyl peptidase from Lactococcus lactis Spp. Lactis: purification and properties, Journal of Applied Microbiology, 68 (4): 357–366.