Probing the Structure of a mutant (Cys131 Asp) of the Human Acidic fibroblast Growth Factor using multidimensional NMR

 

 

Leena P.S.T, Kumar T.K.S and Yu.C

Department of Chemistry, National Tsing Hua University

Hsinchu, Taiwan. R.O.C.

 

Human acidic Fibroblast Growth Factor is a 17 kDa, heparin-binding protein exhibiting a wide array of biological activities.  hFGF-1 is an all b-sheet protein and the heparin-binding segment is postulated to be located in the amino acid segment spanning residues, 120 to 142.  In the present research proposal we aim to study the effects of introducing a negative charge in the heparin-binding site of hFGF-1 by replacing Cys 131 with Aspartic acid using site-directed mutagenesis technique.  Heparin-sepharose affinity profiles obtained over a stepwise NaCl gradient revealed that this Cys131Asp (C131D) mutant has a weak heparin binding affinity.  Preliminary results based on multidimensional NMR techniques showed that introduction of a negative charge in the heparin binding domain triggers drastic changes in the three-dimensional structure of the protein.  Determination of the three-dimensional structure of the C131D mutant of hFGF-1 is expected to provide strong clues on the structural consequences of the mutation C131®D