FOLDED CONFORMATIONS OF ANTIGENIC PEPTIDES FROM RIBOFLAVIN CARRIER PROTEIN IN AQUEOUS HEXAFLUOROACETONE

Riboflavin carrier protein (RCP) plays an important role in transporti ng vitamin B-2 across placental membranes, a process critical for main tenance of pregnancy. Association of the vitamin with the carrier prot ein ensures optimal bioavailability, facilitating transport. The confo rmations of three antigenic peptide fragments encompassing residues 4- 23 (N21), 170-186 (R18), and 200-219 (Y21) from RCP, which have earlie r been studied as potential leads toward a synthetic peptide-based con traceptive vaccine, have been investigated using CD and NMR spectrosco py in aqueous solution and in the presence of the structure-stabilizin g cosolvent hexafluoroacetone trihydrate (HFA). In aqueous solution at pH 3.0, all three peptides ate largely unstructured, with limited hel ical population for the peptides R18 and Y21. The percentage of helici ty estimated from CD experiments is 10% for both the peptides. A drama tic structural transition from an unstructured state to a helical stat e is achieved with addition of HFA, as evidenced by intensification of CD bands at 222 nm and 208 nm for Y21 and R18. The structural transit ion is completed at 50% HFA (v/v) with 40% and 35% helicity for R18 an d Y21, respectively. No structural change is evident for the peptide N 21, even in the presence of HFA. NMR analysis of the three peptides in 50% HFA confirms a helical conformation of Rig and Y21, as is evident from upfield shifts of (CH)-H-alpha resonances and the presence of ma ny sequential NH/NH NOEs with many medium-range NOEs. The helical conf ormation is well established at the center of the sequence, with subst antial fraying at the termini for both the peptides. An extended confo rmation is suggested for the N21 peptide from NMR studies. The helical region of both the peptides (R18, Y21) comprises the core epitopic se quence recognized by the respective monoclonal antibodies. These resul ts shed some light on the issue of structure and folding of antigenic peptides.