Molecular Cloning and Overexpression of WAP Domain of Anosmin-1 (a-WAP) in Escherichia coli

Abstract

Fibroblast Growth Factor (FGF) signaling plays a vital role in a wide range of cellular responses (Detillieux et al., 2003; Freeman et al., 2003). The activation of FGF Receptor (FGFR) is a crucial step in the diverse FGF signaling pathway ( Ayari & Soussi-Yanicostas, 2007). Various intraand extracellular modulators are involved in the FGF signaling pathway. Anosmin-1, an extracellular matrix associated glycosylated protein, is a newly identified modulator of the FGF-mediated signaling process (Bribian et al., 2006; GonzalezMartinez et al., 2004). Anosmin-1 is the product of KAL1 gene. The Kallman syndrome (KS) is a manifestation of the different loss-of-function mutations in the KAL1 gene. The most characteristic features of the Kallman syndrome are anosmia (lack of smell) and hypogonadotrophic hypogonadism (Maestre, 1856; Kallmann et al., 1944). The biological role(s) of Anosmin-1 has been studied both at the cellular and at the biochemical level. Anosmin-1 is suggested to be involved in cell adhesion, cell migration, cell proliferation as well as cell differentiation (Andrenacci et al., 2004; Andrenacci et al., 2006; Ardouin et al., 2000; Bribian et al., 2006; Cariboni et al., 2004; Ernest et al., 2007; Hardelin et al., 1999; Hu et al., 2004; Okubo et al., 2006; Robertson et al., 2001; Rugrali et al., 1996; Soussi-Yanicostas, 1996; Soussi-Yanicostas et al., 1998; Soussi-Yanicostas et al., 2002; Yanicostas et al., 2008;). Anosmin-1 is shown to be a heparin binding protein and formation of the heparin-Anosmin binary complex is believed to be crucial for the function of the protein (Bulow et al., 2002). Interestingly, recent reports suggest a direct interaction between the Nterminus of Anosmin-1 and the FGF-2/FGFR-1/HS ternary complex (Hu et al., 2009; Hu & Bouloux, 2010). The structure of Anosmin-1(Figure 1) comprises of an N-terminal cysteine-rich domain (CR), whey acidic like-protein domain (a-WAP), four fibronectin type III (FnIII) repeats and C-terminal histidine rich region. Recent reports suggest direct interactions of some of the structural domains of Anosmin-1, including a-WAP, with the fibroblast growth factor receptor(s). Although the Anosmin-FGFR binding interface is still not mapped, it is strongly believed that the interactions of Anosmin-1 with FGFR(s) regulates the activation of the receptor.