The interaction between endopolygalacturonase from Fusarium moniliforme and PGIP from Phaseolus vulgaris studied by surface plasmon resonance and mass spectrometry.

Benedetta Mattei,Felice Cervone,Peter Roepstorff

doi:10.1002/cfg.113

Abstract

A combination of surface plasmon resonance (SPR) and matrix-assisted laser-desorptionionization- time-of-flight mass spectrometry (MALDI-TOF-MS) was used to study the interaction between endopolygalacturonase (PG) from Fusarium moniliforme and a polygalacturonase-inhibiting protein (PGIP) from Phaseolus vulgaris. PG hydrolyses the homogalacturonan of the plant cell wall and is considered an important pathogenicity factor of many fungi. PGIP is a specific inhibitor of fungal PGs and is thought to be involved in plant defence against phytopathogenic fungi. SPR was used either to study the effect of the PG glycosylation on the formation of the complex with PGIP, and as a sensitive affinity capture of an interacting peptide from a mixture of PG fragments obtained by limited proteolysis. Mass spectrometry allowed to characterise the interacting peptide eluted from the sensor surface.

Highlights

Surface plasmon resonance (SPR) biosensors are important and versatile tools for studying protein– protein interactions; they allow to measure interactions in real time and require very little material, which usually does not need any chemical modification
surface plasmon resonance (SPR) technology coupled with mass spectrometry can be used to identify and characterise proteins eluted from sensor surfaces (Sonksen et al, 2000; Williams, 2000)
Once a sample containing a mixture of possible ligands is passed over the sensor surface, and binding is detected by the SPR signal, the identification of interacting proteins at the femtomole level is made possible by the use of sensitive mass spectrometers and advanced database searching algorithms (Nelson et al, 2000)

Summary

Introduction

Surface plasmon resonance (SPR) biosensors are important and versatile tools for studying protein– protein interactions; they allow to measure interactions in real time and require very little material, which usually does not need any chemical modification. SPR technology coupled with mass spectrometry can be used to identify and characterise proteins eluted from sensor surfaces (Sonksen et al, 2000; Williams, 2000). Once a sample containing a mixture of possible ligands is passed over the sensor surface, and binding is detected by the SPR signal, the identification of interacting proteins at the femtomole level is made possible by the use of sensitive mass spectrometers and advanced database searching algorithms (Nelson et al, 2000). Polygalacturonase-inhibiting proteins (PGIPs), present in the cell wall of many plants, form specific complexes with fungal PGs and favour the accumulation of oligogalacturonides able to elicit plant defence responses (Cervone et al, 1997). We Copyright # 2001 John Wiley & Sons, Ltd

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