A new chitinase-like xylanase inhibitor protein (XIP) from coffee (Coffea arabica) affects Soybean Asian rust (Phakopsora pachyrhizi) spore germination

Erico Ar Vasconcelos,Claudia V Godoy,Osmundo B Oliveira-Neto,Edivaldo Xf Filho,Marilia S Silva,Daniel Price,Maria F Grossi-De-Sa,John A Gatehouse,Claudine Ds Seixas,Celso G Santana,Leonora Rs Moreira,Elaine Fitches,Angela Mehta

doi:10.1186/1472-6750-11-14

Erico Ar Vasconcelos, Claudia V Godoy + Show 11 more

Open Access

https://doi.org/10.1186/1472-6750-11-14

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Abstract

BackgroundAsian rust (Phakopsora pachyrhizi) is a common disease in Brazilian soybean fields and it is difficult to control. To identify a biochemical candidate with potential to combat this disease, a new chitinase-like xylanase inhibitor protein (XIP) from coffee (Coffea arabica) (CaclXIP) leaves was cloned into the pGAPZα-B vector for expression in Pichia pastoris.ResultsA cDNA encoding a chitinase-like xylanase inhibitor protein (XIP) from coffee (Coffea arabica) (CaclXIP), was isolated from leaves. The amino acid sequence predicts a (β/α)8 topology common to Class III Chitinases (glycoside hydrolase family 18 proteins; GH18), and shares similarity with other GH18 members, although it lacks the glutamic acid residue essential for catalysis, which is replaced by glutamine. CaclXIP was expressed as a recombinant protein in Pichia pastoris. Enzymatic assay showed that purified recombinant CaclXIP had only residual chitinolytic activity. However, it inhibited xylanases from Acrophialophora nainiana by approx. 60% when present at 12:1 (w/w) enzyme:inhibitor ratio. Additionally, CaclXIP at 1.5 μg/μL inhibited the germination of spores of Phakopsora pachyrhizi by 45%.ConclusionsOur data suggests that CaclXIP belongs to a class of naturally inactive chitinases that have evolved to act in plant cell defence as xylanase inhibitors. Its role on inhibiting germination of fungal spores makes it an eligible candidate gene for the control of Asian rust.

Highlights

Asian rust (Phakopsora pachyrhizi) is a common disease in Brazilian soybean fields and it is difficult to control
In this paper we report cloning, heterologous expression and enzymatic features of a new chitinase-like xylanase inhibitor protein (XIP) from coffee (Coffea arabica) (CaclXIP - Coffea arabica Chitinase-like Xylanase Inhibitor Protein), originally identified in the coffee genome [9] as a Class III Chitinase
Heterologous expression and purification of CaclXIP Analysis of sequences present in the Coffee Genome Data Bank identified a type III chitinase-like gene, present in contig 14550, which codes to a xylanase inhibitor protein

Summary

Introduction

Asian rust (Phakopsora pachyrhizi) is a common disease in Brazilian soybean fields and it is difficult to control. Xylanase inhibitor proteins (XIP) are potential “defence” molecules, which could act to prevent plant cell wall degradation by fungal hydrolytic enzymes. They have sequence similarity to glycoside hydrolases of family 18 (GH18) that are plant class III chitinases (PR8). The GH18 family includes naturally inactive chitinases showing (b/a) topology that are predicted to show no catalytic activity due to mutations in the catalytic domain. Some of these proteins have been identified as inhibitors of xylanases (belonging to glycoside hydrolase families GH10 and GH11). Structural features of these (b/a) chitinase-like xylanase inhibitors, as well its interaction with xylanases, has been reviewed recently [2]

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