Poster: Cell Walls

Abs # 1246: Isolation and characterization of an extensin precursor from tobacco

Presenter:	Terneus, Kimberly , BioKat1@aol.com
Authors	Terneus, Kimberly  (A)   Kieliszewski, Marcia J (A)   Leyham, Joseph F (B)
Affiliations:	(A): Department Chemistry and Biochemistry Ohio University (B): Macromolecular Biology Gacility Michigan State University

Extensins are members of the hydroxyproline-rich glycoprotein (HRGP) superfamily and are major components contributing to primary cell wall architecture. They form porous covalently cross-linked networks in the wall during normal cell growth and in response to pathogens and stress. We have purified and biochemically characterized an abundant HRGP salt eluted from the cell surface of tobacco cell suspension cultures (Bright Yellow 2) . The protein is about 40% carbohydrate on a dry weight basis, the main saccharides being arabinose and galactose. A hydroxyproline arabinoside (Hyp-Ara) profile revealed the protein to be mainly Hyp-Ara₄, with lesser amounts of Hyp-Ara₃ Hyp-Ara₂, Hyp-Ara₁, and little nonglycosylated Hyp in small amounts. Isodityrosine (IDT) assays and cross-linking assays demonstrated that the protein contains small amount IDT and has the ability to crosslink. Amino acid composition revealed the protein to consist mainly of hydroxyproline, tyrosine, and lysine with smaller amount of threonine, serine, proline, valine histidine and alanine. Peptide mapping and sequence analysis of the major peptides yielded the following sequences: Ser-Hyp-Hyp-Hyp-Thr-Hyp-Val-Tyr-Lys and Lys-Pro-Tyr-Tyr-Pro-Hyp-His-Thr-Hyp-Val-Tyr-Lys and identified the protein as a P1 type cross-linking extensins. This glycoprotein corresponds to tobacco cDNA clone pCNT1 whose transcript is abundant in tobacco roots, stems, and leaves.