title: Understanding the role of defective invertases in plants: tobacco Nin88 fails to degrade sucrose.
abstract: Cell wall invertases (cwINVs), with a high affinity for the cell wall, are fundamental enzymes in the control of plant growth, development, and carbon partitioning. Most interestingly, defective cwINVs have been described in several plant species. Their highly attenuated sucrose (Suc)-hydrolyzing capacity is due to the absence of aspartate-239 (Asp-239) and tryptophan-47 (Trp-47) homologs, crucial players for stable binding in the active site and subsequent hydrolysis. However, so far, the precise roles of such defective cwINVs remain unclear. In this paper, we report on the functional characterization of tobacco (Nicotiana tabacum) Nin88, a presumed fully active cwINV playing a crucial role during pollen development. It is demonstrated here that Nin88, lacking both Asp-239 and Trp-47 homologs, has no invertase activity. This was further supported by modeling studies and site-directed mutagenesis experiments, introducing both Asp-239 and Trp-47 homologs, leading to an enzyme with a distinct Suc-hydrolyzing capacity. In vitro experiments suggest that the addition of Nin88 counteracts the unproductive and rather aspecific binding of tobacco cwINV1 to the wall, leading to higher activities in the presence of Suc and a more efficient interaction with its cell wall inhibitor. A working model is presented based on these findings, allowing speculation on the putative role of Nin88 in muro. The results presented in this work are an important first step toward unraveling the specific roles of plant defective cwINVs.
authors: Laboratory of Molecular Plant Biology, Katholieke Universiteit Leuven, B-3001 Leuven, Belgium.
authors: Katrien
authors: K
authors: Le Roy
authors: Rudy
authors: R
authors: Vergauwen
authors: Tom
authors: T
authors: Struyf
authors: Shuguang
authors: S
authors: Yuan
authors: Willem
authors: W
authors: Lammens
authors: Janka
authors: J
authors: Mátrai
authors: Marc
authors: M
authors: De Maeyer
authors: Wim
authors: W
authors: Van den Ende
mesh_terms: D000595:Amino Acid Sequence; D055162:Biocatalysis; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D010843:Pichia / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity; D013395:Sucrose / Q000378:metabolism*; D014026:Nicotiana / Q000201:enzymology*; D043324:beta-Fructofuranosidase / Q000737:chemistry / Q000378:metabolism*
chemical_list: D018076:DNA, Complementary; D050505:Mutant Proteins; D010940:Plant Proteins; D011994:Recombinant Proteins; D013395:Sucrose; D043324:beta-Fructofuranosidase