Aging of Dry Desiccation-Tolerant Pollen Does Not Affect Protein Secondary Structure

Abstract

Protein secondary structure and membrane phase behavior i n aging Jypha latifolia pollen were studied by means of Fourier transform infrared microspectroscopy (FTIR). Membranes isolated from fresh pollen occurred mainly in the liquid crystalline phase at room temperature, whereas the membrane fluidity of aged pollen was drastically decreased. This decrease did not result i n large-scale irreversible protein aggregation, as was concluded from i n situ FTIR assessment of the amide-1 bands. Curve-fitting on the infrared absorbance spectra enabled estimation of the proportion of different classes of protein secondary structure. Membrane proteins had a relatively large amount of a-helical structure (48%; band at 1658 cm-'1, and turn-like structures (at 1637 and 1680 cm-') were also detected. The secondary protein structure of isolated cytoplasmic proteins resembled that of proteins in whole pollen and was conserved upon drying in the absence of sucrose. The isolated cytoplasmic proteins had a large amount of a-helical structure (43%), and also P-sheet (at 1637 and 1692 cm-') and turn structures were detected. Heat-denaturing experiments with intact hydrated pollen showed low (1627 cm-') and high (1692 cm-') wave number bands indicating irreversible protein aggregates. The results presented in this paper show that FTIR is an extremely suitable technique to study protein secondary structure in intact plant cells of different hydration levels and developmental stages.Cattail (Typka latifolia) pollen can be dried to water contents of less than 0.05 g H,O g-' dry weight without losing viability. In an atmosphere of 40% RH the pollen attains about 0.06 g H,O g-' dry weight and has a maximum life span of approximately 120 d at 24°C in the dark (van Bilsen and Hoekstra, 1993). At the higher RH of 75% the water content increases to approximately 0.20 g H,O g-' dry

Conformation of a Group 2 Late Embryogenesis Abundant Protein from Soybean. Evidence of Poly (l-Proline)-type II Structure

“…Although group 1 and 2 LEA proteins have a much lower predicted ␣-helical content than group 3 LEA proteins, increases in ␣-helical content arising from changes in ion or sugar concentrations or hydration status could contribute to functional interactions with other biomolecules. ␣-Helical structures were found to compose approximately 40% of the overall protein secondary structures in dry cattail pollen (Wolkers and Hoekstra, 1995). Similar ␣-helical contributions to overall protein secondary structure have also been observed in maize embryos (Wolkers et al, 1998a) and carrot (Daucus carota) somatic embryos (Wolkers et al, 1998b) consistent with earlier predictions of a functional contribution of ␣-helical conformations by several LEA proteins (Dure et al, 1989;Dure, 1993).…”

Fourier Transform Infrared Microspectroscopy Detects Changes in Protein Secondary Structure Associated with Desiccation Tolerance in Developing Maize Embryos1

“…The bands at 2928 and 2856 cm Ϫ1 represent C-H stretching vibrations, arising mainly from neutral lipids, proteins, and carbohydrates. In the 1700 to 1500 cm Ϫ1 region, the amide-I band around 1650 cm Ϫ1 and the amide-II band around 1550 cm Ϫ1 can be observed, which are attributable to proteins (Wolkers and Hoekstra, 1995). The band around 1740 cm Ϫ1 in this region is attributable to ester bonds arising from lipids.…”

“…FTIR spectra were recorded on a spectrometer (model 1725, Perkin-Elmer) equipped with a liquid nitrogencooled mercury/cadmium/telluride detector and a PerkinElmer microscope as described previously (Wolkers and Hoekstra, 1995).…”

“…Differences in the CϭO stretching vibrations of the peptide groups (the amide-I region between 1600-1700 cm Ϫ1 ) provide information on the type of secondary structure, such as ␣-helix, ␤-strands, and different kinds of turn structures. In situ FTIR has been recently applied to study the overall protein secondary structure in dry pollen (Wolkers and Hoekstra, 1995) and seeds (Golovina et al, 1997b). The advantage of FTIR is that protein secondary structure is measured in the native environment of the proteins and that it is a noninvasive technique.…”

Environmental pollution changes in membrane lipids, antioxidants and vitality of Scots pine (Pinus sylvestris L.) pollen

“…However, in these studies no explanation of the phenomenon of pollen injuries was proposed. Investigations of Hoekstra et al (1992), Wolkers and Hoekstra (1995) and Golovina et al (1998) showed that cytoplasmic membranes were prime targets for destructive changes in pollen grains as a result of abiotic stresses. There is little information in the literature about the structural and biophysical function of membranes in the tolerance of pollen grains to industrial pollution.…”