RESEARCH ARTICLE


Functional and Structural Characterization of a Novel Isoamylase from Ostreococcus tauri and Role of the N-Terminal Domain



Nicolas Hedín1, Julieta Barchiesi1, Diego F. Gomez-Casati1, *, María V. Busi1, *
1 Centro de Estudios Fotosintéticos y Bioquímicos (CEFOBI), CONICET - Universidad Nacional de Rosario, Suipacha 531, 2000 Rosario, Argentina


© 2020 Hedín et al

open-access license: This is an open access article distributed under the terms of the Creative Commons Attribution 4.0 International Public License (CC-BY 4.0), a copy of which is available at: (https://creativecommons.org/licenses/by/4.0/legalcode). This license permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

* Address correspondence to this author at the Centro de Estudios Fotosintéticos y Bioquímicos (CEFOBI), CONICET - Universidad Nacional de Rosario, Suipacha 531, 2000 Rosario, Argentina; Fax: +54-0341-4370044; Tel: +54-0341-4371955; E-mails: busi@cefobi-conicet.gov.ar, gomezcasati@cefobi-conicet.gov.ar


Abstract

Background:

The debranching starch enzymes, isoamylase 1 and 2 are well-conserved enzymes present in almost all the photosynthetic organisms. These enzymes are involved in the crystallization process of starch and are key components which remove misplaced α-1,6 ramifications on the final molecule.

Aim:

In this work, we performed a functional and structural study of a novel isoamylase from Ostreococcus tauri.

Methods:

We identified conserved amino acid residues possibly involved in catalysis. We also identified a region at the N-terminal end that resembles a Carbohydrate Binding Domain (CBM), which is more related to the family CBM48, but has no spatial conservation of the residues involved in carbohydrate binding.

Results:

The cloning, expression and biochemical characterization of this N-terminal region confirmed that it binds to polysaccharides, showing greater capacity for binding to amylopectin rather than total starch or amylose.

Conclusion:

This module could be a variant of the CBM48 family or it could be classified within a new CBM family.

Keywords: Carbohydrate Binding Domain (CBM), Debranching starch enzymes, Amino acid residues, Polysaccharides, Biochemical characterization, Carbohydrates.