Combining acetic acid extraction and high-performance gel chromatography in guanidine HCl, extensin secreted into the medium by tobacco (Nicotiana tabacumL. varXanthi) culture cells was separated into three component molecules, namely a major 74-kDa, and two minor 45-and 28-kDa components, in addition to larger oligomers. The sizes of these native extensin molecules were first reasonably assessed using this gel-chromatography system. After deglycosylation with hydrogen fluoride, the separation was improved and the estimated molecular sizes were reduced to 52 kDa, 34 kDa and 18 kDa, respectively. The amino acid compositions of these components were similar, and N-terminal sequences of the 52- and 34-kDa components coincided. The relative abundance of the components was as follows: oligomers, 46; 52-kDa, 44; 34-kDa, 7.7; 18-kDa, 2.2; respectively, on a protein basis (w/w). Fluorography of the acid extract of microsomes from cells labelled with14C-proline revealed only one precursor band of 110-kDa or 42-kDa under the glycosylating or non-glycosylating conditions, respectively. The smaller components in the medium may be derived, by proteolytic cleavage, from the major extensin molecule after secretion.
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