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Although considerable structural information can be acquired from studies carried out on specific domains from IGFBP 1 6, the three dimensional structures of full length IGFBPs haven't yet been determined. A crucial challenge in the structural characterization of full length IGFBPs has been the difficulty in expressing these proteins at levels suitable for NMR/X ray crystallography Lapatinib analysis. We recently reported the first high-yield term and structural characterization of full-length recombinant human IGFBP 2 in E. coli. This opens up new paths to carry out framework based studies in this protein family. Table 2 provides the high res 3D structures received thus far for the individual domains from various IGFBPs and their complexes with IGF 1 using NMR or X ray crystallography. Based on their disulfide bonding routine and houses, the IGFBPs are considered to be thyroglobulin form 1 domain homologues. Both the N terminal and the Cterminal domains are of /B type comprising 30?40% of deposits in normal secondary structural elements and 60?70% in unstructured locations. Figure 2 represents the 3D structure of the N terminal Lymphatic system domain of IGFBP 4 and the C terminal domain of IGFBP 2 based on NMR and X ray crystallography, respectively. Also shown is just a ternary complex relating to the C and N terminal domains of IGFBP 4 and IGF 1. The central or linker domain in all IGFBPs is basically unstructured and contains sites of proteolysis and post translational modification. Reports involving site directed mutagenesis have identified critical residues in IGFBPs that are required for binding the IGFs. These studies also have revealed that both the C terminal domains and N in IGFBPs are crucial for IGF 1/2 binding. It has been proven that truncated IGFBP molecules lacking the N or C terminal domains have considerably paid off binding affinity for the IGFs compared to the full length protein. One particular study within our laboratories centered on the binding affinities JZL184 of truncation mutants of IGFBP 2 for IGF 1. This research has provided valuable insights into IGF binding and is briefly discussed below. Investigation of IGF 1 binding by IGFBP 2 truncation mutants All six IGFBPs contain a CWCV theme within their C terminal domain. In a previous study involving IGFBP 2, derivatives both up and down stream of this motif were identified to be involved in IGF 1 binding. Ergo to check the contribution of the polypeptide part of downstream of the CWCV motif in IGFBP 2, three different truncation mutants comprising elements 1?190, 1?248 and 249?289 were cloned and expressed in E. coli. Even though large, IGFBP 21?248 showed a 20 fold decrease in binding affinity compared to the full length IGFBP 2 and IGFBP 21?190 had a binding affinity indistinguishable from that of IGFBP 21?248. A significant result was borne out by kinetic studies, which unveiled that dissociation of bound IGF 1 from IGFBP 21?248 was considerably faster compared to full length protein.