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Title 

Mutational analysis of Xanthomonas harpin HpaG identifies a key functional region that elicits the hypersensitive response in nonhost plants

Authors 

J G KimE K JeonJ H OhMoon Jae SunI G Hwang

Publisher 

American Society for Microbiology

Issue Date 

2004

Citation 

Journal of Bacteriology, vol. 186, no. 18, pp. 6239-6247

Keywords 

bacterial proteinhydroxylamineprotein hpagprotein xopAunclassified drugalpha helixamino acid analysisconcentration (parameters)erwiniagene identification

Abstract 

HpaG is a type III-secreted elicitor protein of Xanthomonas axonopodis pv. glycines. We have determined the critical amino acid residues important for hypersensitive response (HR) elicitation by random and site-directed mutagenesis of HpaG and its homolog XopA. A plasmid clone carrying hpaG was mutagenized by site-directed mutagenesis, hydroxylamine mutagenesis, and error-prone PCR. A total of 52 mutants were obtained, including 51 single missense mutants and 1 double missense mutant. The HR elicitation activity was abolished in the two missense mutants [HpaG(L50P) and HpaG(L43P/L50P)]. Seven single missense mutants showed reduced activity, and the HR elicitation activity of the rest of the mutants was similar to that of wild-type HpaG. Mutational and deletion analyses narrowed the region essential for elicitor activity to the 23-amino-acid peptide (H2N-NQGISEKQLDQLLTQLIMALLQQ-COOH). A synthetic peptide of this sequence possessed HR elicitor activity at the same concentration as the HpaG protein. This region has 78 and 74% homology with 23- and 27-amino-acid regions of the HrpW harpin domains, respectively, from Pseudomonas and Erwinia spp. The secondary structure of the peptide is predicted to be an α-helix, as is the HrpW region that is homologous to HpaG. The predicted α-helix of HpaG is probably critical for the elicitation of the HR in tobacco plants. In addition, mutagenesis of a xopA gene yielded two gain-of-function mutants: XopA(F48L) and XopA(F48L/M52L). These results indicate that the 12 amino acid residues between L39 and L50 of HpaG have critical roles in HR elicitation in tobacco plants.

ISSN 

0021-9193

Link 

http://dx.doi.org/10.1128/JB.186.18.6239-6247.2004

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2017-04-19


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