Elucidation of different cold-adapted Atlantic cod (Gadus morhua) trypsin X isoenzymes

• Published in: Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics
• Main Authors: Stefánsson, Bjarki, Sandholt, Gunnar Birgir, Guðmundsdóttir, Ágústa
• Other Authors: Matvæla- og næringarfræðideild (HÍ), Faculty of Food Science and Nutrition (UI), Heilbrigðisvísindasvið (HÍ), School of Health Sciences (UI), Háskóli Íslands, University of Iceland
• Format: Article in Journal/Newspaper
• Language: English
• Published: Elsevier BV 2017
• Subjects: Biophysics; Analytical Chemistry; Biochemistry; Molecular Biology; Lífefnafræði; Efnagreining; Lífeðlisfræði; Sameindalíffræði; Biomedicine; Lífvísindi; Atlantic cod; Atlantshafslax; atlantic cod; Gadus morhua; Iceland
• Online Access: https://hdl.handle.net/20.500.11815/153; https://doi.org/10.1016/j.bbapap.2016.10.005

Trypsins from Atlantic cod (Gadus morhua), consisting of several isoenzymes, are highly active cold-adapted serine proteases. These trypsins are isolated for biomedical use in an eco-friendly manner from underutilized seafood by-products. Our group has explored the biochemical properties of trypsins and their high potential in biomedicine. For broader utilization of cod trypsins, further characterization of biochemical properties of the individual cod trypsin isoenzymes is of importance. For that purpose, a benzamidine purified trypsin isolate from Atlantic cod was analyzed. Anion exchange chromatography revealed eight peaks containing proteins around 24 kDa with tryptic activity. Based on mass spectrometric analysis, one isoenzyme gave the best match to cod trypsin I and six isoenzymes gave the best match to cod trypsin X. Amino terminal sequencing of two of these six trypsin isoenzymes showed identity to cod trypsin X. Three sequence variants of trypsin X were identified by cDNA analysis demonstrating that various forms of this enzyme exist. One trypsin X isoenzyme was selected for further characterization based on abundance and stability. Stepwise increase in catalytic efficiency (kcat/Km) of this trypsin X isoenzyme was obtained with substrates containing one to three amino acid residues. The study demonstrates that the catalytic efficiency of this trypsin X isoenzyme is comparable to that of cod trypsin I, the most abundant and highly active isoenzyme in the benzamidine cod trypsin isolate. Differences in pH stability and sensitivity to inhibitors of the trypsin X isoenzyme compared to cod trypsin I were detected that may be important for practical use. Research funded by AVS R&D Fund of Ministry of Fisheries and Agriculture in Iceland (R069-08, R11 028-11, R14 044-14) | Science and Technology Development Fund (120852-0611, 131804-0611) Ritrýnt tímarit Peer Reviewed Pre-Print