Yt. Zhou et al., Genetically directed synthesis and spectroscopic analysis of a protein polymer derived from a flagelliform silk sequence, BIOMACROMOL, 2(1), 2001, pp. 111-125
The flagelliform silk protein underlies the unique elastomeric properties d
isplayed by the capture spiral of arachnid webs. To investigate molecular m
echanism underlying the elastomeric recovery of the capture spiral, a model
polypeptide based upon the elastomeric repeat sequence of Nephila clavipes
flagelliform silk protein has been synthesized using recombinant DNA techn
iques. Polypeptide 1 contains 11 repeats of the 25 amino acid sequence [(Gl
y-Pro-Gly-Gly-Ser-Gly-Pro-Gly-Gly-Tyr)(2)-Gly-Pro-Gly-Gly-Lys] and was expr
essed in Escherichia coil strain BL21(DE3) as a C-terminal fusion to a deca
histidine leader sequence. A combination of H-1-H-1 COSY, DEPT,H-1-C-13 HET
COR, and H-1-C-13 HMBC NMR spectroscopy was employed on polypeptides 1 and
the [1-C-13]glycine-labeled analogue 1G to assign the H-1 and C-13 NMR reso
nances of the amino acid residues comprising the flagelliform silk repeat s
equence. The conformational properties of 1 in aqueous solution were invest
igated using a combination of CD, FTIR, VT-NMR, and two-dimensional NOESY N
MR. These techniques were consistent with the presence of small but detecta
ble population of beta -turn conformers between Gly(1) and Gly(4) of the pe
ntapeptide units of 1. FTIR and CD studies of solid films of 1 indicated an
increase in beta -turn population in the solid state, which coincided with
the decrease in hydration level of the polypeptide. The spectroscopic info
rmation suggests that the pentapeptide segments of the flagelliform silk pr
otein adopt a beta -turn conformation in the fiber and that the mechanism o
f elasticity may resemble that proposed for other beta -turn forming polype
ptides including elastin.