Purification and characterization of proline/hydroxyproline-rich glycoprotein from pearl millet coleoptiles infected with downy mildew pathogen Sclerospora graminicola.
Hydroxyproline-rich glycoproteins (HRGPs) are important plant cell wall structural components, which are also involved in response to pathogen attack. In pearl millet, deposition and cross-linking of HRGPs in plant cell walls was shown to contribute to the formation of resistance barriers against the phytopathogenic oomycete Sclerospora graminicola. In the present study, the purification and characterization of HRGPs that accumulated in coleoptiles of pearl millet seedlings in response to S. graminicola inoculation has been carried out. Periodic acid Schiff's staining revealed that the purified protein was a glycoprotein. The protein to carbohydrate ratio was determined to be 95.5%:4.5% (w/w). Proline amounted for 20 mol% of the total amino acids as indicated by amino acid composition analysis. The isolated protein had a pI of 9.8 and was shown to be composed of subunits of 27, 17, and 14 kDa. Cross reactivity with the monoclonal antibody MAC 265 and the presence of the signature amino acid sequence, PVYK, strongly suggested to classify the purified glycoprotein as a member of the P/HRGPs class. In the presence of horseradish peroxidase and H2O2 the purified glycoprotein served as a substrate for oxidative cross-linking processes.
Deepak
Shantharaj
S
Department of Studies in Applied Botany and Biotechnology, University of Mysore, Manasagangotri, Mysore 570006, Karnataka, India.
Shailasree
Sekhar
S
Sujeeth
Neerakkal
N
Kini
Ramachandra K
RK
Shetty
Shekar H
SH
Mithöfer
Axel
A
D002852:Chromatography, Ion Exchange; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D005658:Fungi / Q000737:chemistry* / Q000302:isolation & purification; D006909:Hydroxyproline / Q000737:chemistry*; D013058:Mass Spectrometry; D008897:Panicum / Q000737:chemistry* / Q000382:microbiology; D010455:Peptides / Q000737:chemistry / Q000302:isolation & purification*; D055232:Proline-Rich Protein Domains
D010455:Peptides; D006909:Hydroxyproline