Old and new ways to probe plant cell-wall architecture

Abstract

Wall structure has been analysed by a process of careful demolition, in which chemical extradants are used to remove specific polymers for sugar and linkage analysis, gel-permeation or ion-exchange chromatography, and nuclear magnetic resonance spectroscopy. Sequence-dependent endoglycanases cleave certain polysaccharides into oligomers that can be sequenced completely and give a clear picture of the repetitive units used to make fundamental polymers. We have also developed and adapted new chemical procedures and pulse-labelling techniques to give more information on the ways that wall polymers are subtly modified during growth. In this report, we review these conventional means of carbohydrate analyses together with newer methods of selective enzymic hydrolysis, separation of large oligosaccharides by high pH anion-exchange chromatography, and detection of molecular mass of several thousand daltons by electrospray mass spectrometry. These new technologies have already given much valuable information about the polymeric building blocks, but little information on how these polymers are arranged in space. For this, we adapted new cryopreservation techniques for electron microscopy that can image the wall in as close to the in vivo state as possible. In addition to defining anomeric linkages and linkage structures in preparations of native polymers, nuclear magnetic resonance spectroscopy can also determine the relative mobility of particular polymers within the structure of hydrated cell walls. The generation of antibodies to particular cell wall epitopes has enabled us to define architectural differences among species, among tissue types, and even among domains within a single wall. Our awareness of the diversity and complexity of primary cell wall architecture has driven a search for methodologies such as Fourier transform infrared and Fourier transform Raman microspectroscopies, which are suitable for analysis at the single cell wall level. Key words: cell walls, polysaccharides, gas – liquid chromatography – mass spectrometry, Fourier transform infrared microspectroscopy, nuclear magnetic resonance spectroscopy, immunocytochemistry.