Analysis of the expression, subcellular and tissue localisation of phosphoglucan, water dikinase (PWD/GWD3) in Solanum tuberosum L.: a bioinformatics approach for the comparative analysis of two α-glucan, water dikinases (GWDs) from Solanum tuberosum L.

Abstract

Analysis of the expression, subcellular and tissue localisation of phosphoglucan, water dikinase (PWD/GWD3) in Solanum tuberosum L.: a bioinformatics approach for the comparative analysis of two a-glucan, water dikinases (GWDs) from Solanum tuberosum L. Abstract There are several important factors affecting the rate of starch decomposition in plants, including the circadian clock, the regulation of gene expression, the regulation of enzyme activities and starch phosphorylation by glucan, water dikinase activities (GWDs). One isoform of glucan, water dikinase named GWD3 or PWD (EC 2.7.9.5) was isolated for the first time from Arabidopsis thaliana, and now we report its isolation and identification in Solanum tuberosum L. leaves and tubers. We compare StGWD3 sequence to the other GWDs sequences using bioinformatics tools and propose also structural models for the starch-binding domains in StGWD3 and StGWD1. The StGWD3 gene expression and protein were localised in different heterotrophic and autotrophic potato tissues and organs using in situ RT-PCR and immunolocalisation methods, respectively. Diurnal changes in the transcript abundance of StGWD3 in leaves were analysed using quantitative real-time PCR and they appeared to be typical for most genes involved in starch degradation in chloroplasts.Keywords Starch degradation Á Cold-sweetening Á Potato Electronic supplementary material The online version of this article

Non‐GMO potato lines with an altered starch biosynthesis pathway confer increased‐amylose and resistant starch properties

“…Furthermore, there were several proteins present in the IAm starch granules, which were not present in the control. One band was found at approximately 135 kDa corresponding to either SSIII (139 kDa) or PWD/GWD3 (132 kDa) . The other polypeptides present in IAm lines were identified by N‐terminal sequencing.…”

The starch‐binding domain family CBM41—An in silico analysis of evolutionary relationships

“…With regard to the location of an SBD in the domain organization of a polypeptide chain, it may either precede or succeed the catalytic domain . It moreover is frequently found accompanied by so‐called fibronectin type III domains occurring in a wide variety of carbohydrate‐active enzymes .…”

“…SBDs were originally recognized as a module found typically in microbial amylolytic and related enzymes. It is worth mentioning that various SBDs and GBDs have also been identified in other plant and animal enzymes and proteins, such as plant starch synthase III, glucan‐water dikinases, starch‐excess protein‐4, animal laforin, genethonin‐1, as well as the β‐subunit of AMP‐activated protein kinase and its homologues in plants, and even in lytic polysaccharide monooxygenases from fungi . Nevertheless, as for the microbial SBDs, generally the non‐microbial SBDs or GBDs are always involved in metabolism of starch and/or glycogen, or other α‐glucans …”

Two structurally related starch-binding domain families CBM25 and CBM26

“…Although typical SBDs with their main role to enable the amylolytic enzyme to cope with raw starch are associated mainly with microbial amylases (RodriguezSanoja et al 2005a;Machovic & Janecek 2006b), in plants and animals, remarkably, their homologues have adopted various related functions, e.g. in phosphoglucan, water dikinase-3 (Mikkelsen et al 2006;Glaring et al 2011, Orzechowski et al 2013), starch-excess protein-4 (Vander Kooi et al 2010, laforin (Minassian et al 2000;Gentry et al 2013), genethonin-1 (Janecek 2002;Jiang et al 2010) and β-subunit of AMP-activated protein kinase (Hudson et al 2003;Polekhina et al 2003Polekhina et al , 2005. Nevertheless, all these nonmicrobial SBDs (or glycogen-binding domains) have always something to do with α-glucans (Janecek et al 2011).…”