A two-phase model for the non-processive biosynthesis of homogalacturonan polysaccharides by the GAUT1:GAUT7 complex

Amos, Robert A.; Pattathil, Sivakumar; Yang, Jeong-Yeh; Atmodjo, Melani A.; Urbanowicz, Breeanna R.; Moremen, Kelley W.; Mohnen, Debra

doi:10.1074/jbc.RA118.004463

Title: A two-phase model for the non-processive biosynthesis of homogalacturonan polysaccharides by the GAUT1:GAUT7 complex

Journal Article · Sat Dec 01 00:00:00 EST 2018 · Journal of Biological Chemistry

DOI:https://doi.org/10.1074/jbc.RA118.004463· OSTI ID:1771357

Amos, Robert A.; Pattathil, Sivakumar; Yang, Jeong-Yeh; Atmodjo, Melani A.; Urbanowicz, Breeanna R.; Moremen, Kelley W.;

Homogalacturonan (HG) is a pectic glycan in the plant cell wall that contributes to plant growth and development and cell wall structure and function, and interacts with other glycans and proteoglycans in the wall. HG is synthesized by the galacturonosyltransferase (GAUT) gene family. Two members of this family, GAUT1 and GAUT7, form a heteromeric enzyme complex in Arabidopsis thaliana. Here, we established a heterologous GAUT expression system in HEK293 cells and show that co-expression of recombinant GAUT1 with GAUT7 results in the production of a soluble GAUT1:GAUT7 complex that catalyzes elongation of HG products in vitro. The reaction rates, progress curves, and product distributions exhibited major differences dependent upon small changes in the degree of polymerization (DP) of the oligosaccharide acceptor. GAUT1:GAUT7 displayed >45-fold increased catalytic efficiency with DP11 acceptors relative to DP7 acceptors. Although GAUT1:GAUT7 synthesized high-molecular-weight polymeric HG (>100 kDa) in a substrate concentration–dependent manner typical of distributive (nonprocessive) glycosyltransferases with DP11 acceptors, reactions primed with short-chain acceptors resulted in a bimodal product distribution of glycan products that has previously been reported as evidence for a processive model of GT elongation. As an alternative to the processive glycosyltransfer model, a two-phase distributive elongation model is proposed in which a slow phase, which includes the de novo initiation of HG and elongation of short-chain acceptors, is distinguished from a phase of rapid elongation of intermediate- and long-chain acceptors. Upon reaching a critical chain length of DP11, GAUT1:GAUT7 elongates HG to high-molecular-weight products.

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Research Organization:: Univ. of Georgia, Athens, GA (United States)

Sponsoring Organization:: USDOE Office of Science (SC), Basic Energy Sciences (BES). Chemical Sciences, Geosciences, and Biosciences Division; National Science Foundation (NSF); National Institutes of Health (NIH)

Grant/Contract Number:: PS02–06ER64304; SC0015662; FG02–12ER16324; SC0008472; FG02-93ER20097; FG02-12ER16324; DBI-0421683; PS02-06ER64304; 2010-65115-20396; P41GM103390; P01GM107012.

OSTI ID:: 1771357

Alternate ID(s):: OSTI ID: 1611056

Journal Information:: Journal of Biological Chemistry, Journal Name: Journal of Biological Chemistry Vol. 293 Journal Issue: 49; ISSN 0021-9258

Publisher:: ElsevierCopyright Statement

Country of Publication:: United States

Language:: English

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Cited by: 25 works

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59 BASIC BIOLOGICAL SCIENCES
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY
Biochemistry & Molecular Biology
plant cell wall
glycosyltransferase
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enzyme mechanism
processivity
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Title: A two-phase model for the non-processive biosynthesis of homogalacturonan polysaccharides by the GAUT1:GAUT7 complex

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