Characterization of recombinant CDR1, an Arabidopsis aspartic proteinase involved in disease resistance.
The Arabidopsis thaliana constitutive disease resistance 1 (CDR1) gene product is an aspartic proteinase that has been implicated in disease resistance signaling (Xia, Y., Suzuki, H., Borevitz, J., Blount, J., Guo, Z., Patel, K., Dixon, R. A., and Lamb, C. (2004) EMBO J. 23, 980-988). This apoplastic enzyme is a member of the group of "atypical" plant aspartic proteinases. As for other enzymes of this subtype, CDR1 has remained elusive until recently as a result of its unusual properties and localization. Here we report on the heterologous expression and characterization of recombinant CDR1, which displays unique enzymatic properties among plant aspartic proteinases. The highly restricted specificity requirements, insensitivity toward the typical aspartic proteinase inhibitor pepstatin A, an unusually high optimal pH of 6.0-6.5, proteinase activity without irreversible prosegment removal, and dependence of catalytic activity on formation of a homo-dimer are some of the unusual properties observed for recombinant CDR1. These findings unveil a pattern of unprecedented functional complexity for Arabidopsis CDR1 and are consistent with a highly specific and regulated biological function.
Simões
Isaura
I
Departamento de Biologia Molecular e Biotecnologia, Centro de Neurociências e Biologia Celular, Universidade de Coimbra, Apt. 3126, Coimbra 3000, Portugal.
Faro
Rosário
R
Bur
Daniel
D
Faro
Carlos
C
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000201:enzymology* / Q000235:genetics; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D016282:Aspartic Acid Endopeptidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D001665:Binding Sites; D003001:Cloning, Molecular; D016384:Consensus Sequence; D018076:DNA, Complementary; D018744:DNA, Plant / Q000235:genetics / Q000302:isolation & purification; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics; D016650:Fluorescein-5-isothiocyanate; D005456:Fluorescent Dyes; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D057927:Hydrophobic and Hydrophilic Interactions; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D021141:Nucleic Acid Amplification Techniques; D010935:Plant Diseases / Q000235:genetics*; D011485:Protein Binding; D021382:Protein Sorting Signals; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D013696:Temperature
D029681:Arabidopsis Proteins; D018076:DNA, Complementary; D018744:DNA, Plant; D005456:Fluorescent Dyes; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D016282:Aspartic Acid Endopeptidases; C502093:CDR1 protein, Arabidopsis; D016650:Fluorescein-5-isothiocyanate