The calpain domain of the maize DEK1 protein contains the conserved catalytic triad and functions as a cysteine proteinase.
Development of the aleurone layer of maize grains requires the activity of the Defective kernel 1 (Dek1) gene, encoding a predicted 240-kDa membrane-anchored protein with a C terminus similar to animal calpain domain II&III. Three-dimensional modeling shows that DEK1 domain II contains a conserved calpain catalytic triad and that domain II&III has a predicted structure similar to m-calpain. Recombinant DEK1 domain II&III exhibits activity in the caseinolytic assay in the absence of calcium, although the activity is enhanced by calcium. This is in sharp contrast to animal calpains, which require Ca2+ to be active. Bacterially expressed DEK1 domain II does not display caseinolytic activity, suggesting an important role for DEK1 domain III. Mutation of the catalytic Cys residue to Ser leads to a loss of caseinolytic activity of DEK1 domain II&III. Two features of DEK1 calpain may contribute to maintaining the active site triad in an "active" configuration in the absence of Ca2+, both of which are predicted to keep m-calpain domains IIa and IIb apart. First, DEK1 lacks key charged residues in the basic loop of domain II, and secondly, the absence of an acidic loop in domain III, both of which are predicted to be neutralized upon Ca2+ binding. The Dek1 transcript is present in all cell types in developing maize endosperm, suggesting that the activity of the DEK1 calpain is regulated at the post-transcription level. The role of DEK1 in aleurone signaling is discussed.
Wang
Cunxi
C
Pioneer Hi-Bred International, A DuPont Company, Johnston, Iowa 50131, USA.
Barry
Jennifer K
JK
Min
Zhao
Z
Tordsen
Gabrielle
G
Rao
A Gururaj
AG
Olsen
Odd-Arne
OA
D000595:Amino Acid Sequence; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D002154:Calpain / Q000737:chemistry*; D002384:Catalysis; D002462:Cell Membrane / Q000378:metabolism; D002942:Circular Dichroism; D017124:Conserved Sequence; D003545:Cysteine / Q000737:chemistry; D003546:Cysteine Endopeptidases / Q000737:chemistry* / Q000378:metabolism*; D003553:Cystine / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D019992:Databases as Topic; D004492:Edetic Acid / Q000494:pharmacology; D005285:Fermentation; D017403:In Situ Hybridization; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010802:Phylogeny; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D012323:RNA Processing, Post-Transcriptional; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D013997:Time Factors; D014158:Transcription, Genetic; D003313:Zea mays / Q000201:enzymology / Q000378:metabolism*
D018076:DNA, Complementary; D011994:Recombinant Proteins; D003553:Cystine; D004492:Edetic Acid; D002154:Calpain; D003546:Cysteine Endopeptidases; C449000:m-calpain; D003545:Cysteine; D002118:Calcium