High throughput screening of starch structures using carbohydrate microarrays

Abstract

In this study we introduce the starch-recognising carbohydrate binding module family 20 (CBM20) from Aspergillus niger for screening biological variations in starch molecular structure using high throughput carbohydrate microarray technology. Defined linear, branched and phosphorylated maltooligosaccharides, pure starch samples including a variety of different structures with variations in the amylopectin branching pattern, amylose content and phosphate content, enzymatically modified starches and glycogen were included. Using this technique, different important structures, including amylose content and branching degrees could be differentiated in a high throughput fashion. The screening method was validated using transgenic barley grain analysed during development and subjected to germination. Typically, extreme branching or linearity were detected less than normal starch structures. The method offers the potential for rapidly analysing resistant and slowly digested dietary starches.

Understanding Changes in Tomato Cell Walls in Roots and Fruits: The Contribution of Arbuscular Mycorrhizal Colonization

“…Comprehensive Microarray Polymer Profiling (CoMPP) combines the high-throughput capacity of the microarray with the specificity of monoclonal antibodies (mAbs) and carbohydrate-binding modules (CBMs), to characterize plant cell walls glycomic profiles [5,6]. Through this approach, it is possible to study the relative abundance of, and interactions between, hundreds or thousands of molecules simultaneously, using very small volumes of plant extracts.…”

Expression of starch-binding factor CBM20 in barley plastids controls the number of starch granules and the level of CO2 fixation

“…3.2.1.3). This variant of CBM20 has a very strong affinity for starch and α-glucan (Tanackovic et al , 2016). We added an N-terminal transit peptide (TP) from barley granule-bound starch synthase (GBSSIa) as previous studies have shown that this TP effectively targets the transgenic proteins to plastids, and that it is effectively cleaved off during passage of the protein over the plastid envelope membranes (Carciofi et al , 2012 b ; Hebelstrup et al , 2017).…”

“…CBM20 is one of these SBDs, and entries with CBM20 modules are reported from organisms with broad phylogenetic diversity, including archaea, bacteria, and eukaryotes (Christiansen et al , 2009). The CBM20 used in the present study was the 12 kDa starch-binding domain (residues 509–616) of glycoamylase from Aspergillus niger , which was chosen based on a specific and high affinity for starch (Tanackovic et al , 2016). The affinity of CBM20 for starch is also significantly higher than that of the barley starch synthases GBSSI, SSI, SSIIa, SSIIIa, and SSIV (Cuesta-Seijo et al , 2015).…”

“…Some starch synthases and starch-modifying enzymes contain CBMs that allow them to target to starch granules (Pfister and Zeeman, 2016) whilst others depend on indirect interactions through other proteins that contain CBMs (Seung et al , 2015; Pfister and Zeeman, 2016). Glucoamylase from Aspergillus niger contains a domain of the CBM20 family that has a very high affinity for starch (Christiansen et al , 2009; Cuesta-Seijo et al , 2015; Tanackovic et al , 2016), and in this present study we examined whether this module (hereafter termed simply CBM20) may provide a means by which to explore starch–protein interactions in plastids. Our assumption was that since CBM20 has a very strong affinity for starch, it will competitively interfere with the large array of starch–protein interactions in plastids.…”

Isomalto/malto-polysaccharides from starch : Reaction pathways and new characterization strategies

“…However, these chemical and physical methods cannot compete with the pattern recognition that is displayed in nature. Enzymes are by far, the most sophisticated tools available for pattern recognition in polysaccharide molecules Tanackovic et al, 2016). Enzymes are therefore often used instead of, or in combination with, traditional methods for the characterization of polysaccharides.…”