Isolation and characterization of alliinase cDNA clones from garlic (Allium sativum L.) and related species.
cDNA libraries constructed from poly(A)-rich RNA isolated from Allium sativum (garlic), Allium cepa (onion) and Allium ascalonicum (shallot) were screened for cDNA clones encoding the alliinase using colony hybridization. Sequence analysis of the alliinase cDNA clones from different Alliaceae species revealed a high degree of sequence similarity both at the nucleotide and at the amino acid level. Apparently, the alliinases are translated from mRNA species of approximately 2200 nucleotides. The primary translation products are preproproteins which are converted into the mature alliinases following post-translational modifications. In the case of the garlic alliinase, the mRNA encodes a 486-amino-acid polypeptide with a molecular mass of 55,623 Da. Cleavage of the signal peptide (28 amino acids) results in a preprotein which extends 10 amino acids before the first amino acid of the mature protein of 51,451 Da. Southern-blot analysis of genomic DNA has shown that the alliinases are most probably encoded by a family of closely related genes, which is in good agreement with the sequence heterogeneity found between different alliinase cDNA clones of one species.
Van Damme
E J
EJ
Laboratory for Phytopathology and Plant Protection, Katholieke Universiteit Leuven, Belgium.
Smeets
K
K
Torrekens
S
S
Van Leuven
F
F
Peumans
W J
WJ
D000490:Allium / Q000201:enzymology* / Q000235:genetics; D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D013437:Carbon-Sulfur Lyases / Q000235:genetics*; D004247:DNA / Q000235:genetics* / Q000302:isolation & purification; D005737:Garlic / Q000201:enzymology* / Q000235:genetics; D015723:Gene Library; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D010946:Plants, Medicinal; D011061:Poly A / Q000235:genetics / Q000302:isolation & purification; D021382:Protein Sorting Signals / Q000378:metabolism; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid
D015345:Oligonucleotide Probes; D021382:Protein Sorting Signals; D012333:RNA, Messenger; D011061:Poly A; D012313:RNA; D004247:DNA; D013437:Carbon-Sulfur Lyases; C019243:alliin lyase