Attempts to find the function of Pf-1806301 using antibodies
Wannere, Gauri C.
MetadataShow full item record
Pf -180301 (Pf-1955, a putative archaeal Lsm protein, shows sequence similarity to both archaeal Lsm and bacteria Hfq proteins. Sm/Lsm proteins are a large family of RNA-binding proteins that have been identified in archaea, bacteria and eukarya. The X-ray structure revealed that Pf -1955 exists as dimer of octamer (16 units). The three-dimensional structure of Pf -1955 has revealed structural similarity with Sm/Lsm proteins, but also shows absence of key RNA-binding residues. This thesis attempts to recognize if Pf1955 binds to DNA or RNA. The gene encoding Pf -1955 was recloned into pDESTC1 vector, which provides a Tev cleavage site for the removal of the His-tag after protein purification. The antibodies were generated using purified recombinant protein in solution. However, antibodies failed to recognize Pf -1955 from the native Pyrococcus furiosus cell extract. Any of the several reasons may be attributed to this failure: (a) the concentration of the Pf -1955 in native cell extract is not known (b) the protein should have been submitted as a gel-band to generate specific antibodies oand (c) the 100 C of boiling temperature is not enough to denature Pf -1955.