Low-resolution structural studies of human Stanniocalcin-1

Background: Stanniocalcins (STCs) represent small glycoprotein hormones, found in all vertebrates, which have been functionally implicated in Calcium homeostasis. However, recent data from mammalian systems indicated that they may be also involved in embryogenesis, tumorigenesis and in the context of the latter especially in angiogenesis. Human STCI is a 247 amino acids protein with a predicted molecular mass of 27 kDa, but preliminary data suggested its di- or multimerization. The latter in conjunction with alternative splicing and/or post-translational modification gives rise to forms described as STC(50) and "big STC", which molecular weights range from 56 to 135 kDa. Results: In this study we performed a biochemical and structural analysis of STCI with the aim of obtaining low resolution structural information about the human STCI, since structural information in this protein family is scarce. We expressed STCI in both E. coli and insect cells using the baculo virus system with a C-terminal 6 x His fusion tag. From the latter we obtained reasonable amounts of soluble protein. Circular dichroism analysis showed STCI as a well structured protein with 52% of alpha-helical content. Mass spectroscopy analysis of the recombinant protein allowed to assign the five intramolecular disulfide bridges as well as the dimerization Cys202, thereby confirming the conservation of the disulfide pattern previously described for fish STCI. SAXS data also clearly demonstrated that STCI adopts a dimeric, slightly elongated structure in solution. Conclusion: Our data reveal the first low resolution, structural information for human STCI. Theoretical predictions and circular dichroism spectroscopy both suggested that STCI has a high content of alpha-helices and SAXS experiments revealed that STCI is a dimer of slightly elongated shape in solution. The dimerization was confirmed by mass spectrometry as was the highly conserved disulfide pattern, which is identical to that found in fish STCI