Proteolytic enzymes (known as proteases) catalyze the hydrolytic cleavage of peptide bonds in their target proteins, and about 2% of the genes code for these enzymes in higher organisms [1-3]. Protease inhibitors (PIs) are widely distributed in the plant kingdom and observed in storage tissues, including tubers and seeds [4]. Also, it is thought that these inhibitors play important roles in defense against herbivores that protect against digestion by proteases [5]. Previous studies revealed that the expression of many PI genes is induced by mechani- cal or herbivore damage [6]. PIs have been classified into families and subfamilies based on sequence similarity with protein folds of the inhibitory domains or units. According to their inhibitor domains, PIs can be grouped into 48 families [7]. Soybean trypsin inhibitors (Kunitz), Bowman–Birk inhibitors (BBIs), and potato inhibitors I and II are widely studied classes [8]. BBIs are members of the serine protease inhibitors family and are commonly found in leguminous and cere- al plants [5]. BBIs were first discovered and characterized in soybean [9, 10]. BBIs are double-headed serine pro- teinase inhibitors, having low molecular mass between 8 and 20 kDa and being rich the cysteine residues [11]. These cysteine residues support conservation of the con- formation of the inhibitor with the disulfide link, hence allowing the inhibition of the target enzyme [12]. In dicotyledonous plants, BBIs have a single polypeptide chain with molecular mass of 8 kDa; in contrast, mono- cotyledonous plants contain two groups of BBIs. The first group has a single polypeptide chain with molecular mass of about 8 kDa and a single reactive site. The second group has molecular mass of 16 kDa with two reactive sites [12, 13]. The loops (or reactive sites) contribute to inhibition of trypsin and chymotrypsin in monocotyledo- nous plants and trypsin, chymotrypsin, and elastase in dicotyledonous plants. Also, the three-dimensional struc- ture of peanut, soybean, and barley BBIs showed that there are disulfide bridges between two conserved cysteine residues (e.g. C 1 –C 14 , C 2 –C 6 , C 3 –C 13 , C 4 –C 5 , C 7 –C 9 , C 8 –C 12 , C 10 –C 11 ) [8]. Three major roles were defined as ISSN 0006-2979, Biochemistry (Moscow), 2014, Vol. 79, No. 8, pp. 836-844. © Pleiades Publishing, Ltd., 2014. Published in Russian in Biokhimiya, 2014, Vol. 79, No. 8, pp. 1042-1051. 836 Abbreviations: BBI, Bowman–Birk inhibitor; NCBI, National Center for Biotechnology Information; NJ, neighbor-joining method; PI, protease inhibitor; TMVs, Turkish maize varieties. * To whom correspondence should be addressed. Characterization of Wound-Induced Serine Protease Inhibitor (wip1) Genes and Proteins in Turkish Maize Varieties E. Filiz*, H. Tombuloglu, I. Koc, and E. Osma Duzce University, Department of Crop and Animal Production, Cilimli Vocational School, 81750, Cilimli, Duzce, Turkey; fax: +90-380-6817313; E-mail: ertugrulfiliz@gmail.com Received April 15, 2014 Revision received April 28, 2014 Abstract—Protease inhibitors (PIs) are generally small proteins that have been identified in plants. The wip1 gene codes for wound-induced protein, which is similar to serine PIs of the Bowman–Birk family (BBIs). In this study, we analyzed 10 wip1 genes of Turkish maize varieties to understand the structure and characteristics of the wip1 genes and proteins in maize. We found that genetic variability of wip1 genes was higher (π: 0.0173) than reported in previous studies. Tajima’s D value was found to be positive (1.73), suggesting over-dominant selection in these loci. According to phylogenetic analysis of wip1 pro- teins, monocot and dicot BBIs were separated independently, and Turkish varieties were clustered with each other general- ly. The 3D structures of wip1 proteins indicated that several wip1 proteins had structural divergence in active loops, con- taining various numbers of cysteine residues ranging between 7 and 9. Particularly, Cys74 was identified in Kocbey and Gozdem varieties, whereas Cys98 was only in the Gozdem variety. Also, a critical serine residue (Ser98) was observed in two varieties – Antbey and Batem Efe. These results can contribute to understanding the role of wip1 genes and corresponding proteins in maize. DOI: 10.1134/S0006297914080124 Key words: protease inhibitors, wound-induced protein, Bowman–Birk family, Turkish maize