Spectroscopic characterization and structural modeling of prolamin from maize and pearl millet

Abstract

Biophysical methods and structural modeling techniques have been used to characterize the prolamins from maize ( Zea mays) and pearl millet ( Pennisetum americanum). The alcohol-soluble prolamin from maize, called zein, was extracted using a simple protocol and purified by gel filtration in a 70% ethanol solution. Two protein fractions were purified from seed extracts of pearl millet with molecular weights of 25.5 and 7 kDa, as estimated by SDS-PAGE. The high molecular weight protein corresponds to pennisetin, which has a high alpha-helical content both in solution and the solid state, as demonstrated by circular dichroism and Fourier transform infrared spectra. Fluorescence spectroscopy of both fractions indicated changes in the tryptophan microenvironments with increasing water content of the buffer. Low-resolution envelopes of both fractions were retrieved by ab initio procedures from small-angle X-ray scattering data, which yielded maximum molecular dimensions of about 14 nm and 1 nm for pennisetin and the low molecular weight protein, respectively, and similar values were observed by dynamic light scattering experiments. Furthermore, (1)H nuclear magnetic resonance spectra of zein and pennisetin do not show any signal below 0.9 ppm, which is compatible with more extended solution structures. The molecular models for zein and pennisetin in solution suggest that both proteins have an elongated molecular structure which is approximately a prolate ellipsoid composed of ribbons of folded alpha-helical segments with a length of about 14 nm, resulting in a structure that permits efficient packing within the seed endosperm.

The interaction between heparin and Lys49 phospholipase A2 reveals the natural binding of heparin on the enzyme

“…To evaluate the structural interaction effects induced by heparin on the secondary structure of BthTx-I, we used SOMCD software. This method has been shown to predict secondary structure in proteins very well [12,13]. The spectral analysis indicates that the secondary structure content of the BthTx-I is: 41.7% ␣-helix, 10.4% turn, 13.0% ␤-sheet and 34.9% random coil, which is consistent with the content observed in the crystal structures (PDB accession no.…”

“…3A) was resolved into six bands on the amide I spectral region (1600-1700 cm −1 ). The Gaussian band profiles positions in the amide I region [13][14][15] show the results of this analysis and reveal that the secondary structure content is 43.1% ␣-helix, 15.4% turn, 10.2% ␤-sheet and 31.3% random coil, in close agreement with the secondary structure composition estimated by CD and the crystal structures. A more straightforward interpretation of the interaction of heparin with FTIR should supply results that show how and where the heparin binds on protein without changing the secondary structure content.…”

Small-angle scattering studies of macromolecular solutions

“…anisometry, symmetry) (Petoukhov & Svergun, 2003. DAMMIN has been actively used for practical shape determination by different groups (Aparicio et al, 2002;Arndt et al, 2003;Bugs et al, 2004;Dainese et al, 2005;Egea et al, 2001;Fujisawa et al, 2001;Hammel et al, 2004;Scott et al, 2002). Interestingly, the program also provided meaningful models for non-biological systems with moderate polydispersity (Ozerin et al, 2005;Shtykova et al, 2003).…”

Production of biopolymer composites by particle bonding

“…Several models have been proposed for the 3D shape of α-zein (Argus et al, 1982;Garrett et al, 1993;Tatham et al, 1993;Matsushima et al, 1997;Bugs et al, 2004;Forato et al, 2004). Despite all these trials, a generally accepted axial ratio of the individual molecule has not even been obtained yet.…”