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Molecular identification of tuliposide B‐converting enzyme: a lactone‐forming carboxylesterase from the pollen of tulip
Author(s) -
Nomura Taiji,
Murase Tatsunori,
Ogita Shinjiro,
Kato Yasuo
Publication year - 2015
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1111/tpj.12883
Subject(s) - carboxylesterase , enzyme , pollen , identification (biology) , lactone , biology , chemistry , biochemistry , botany
Summary 6‐Tuliposides A (PosA) and B (PosB), which are the major secondary metabolites in tulip ( Tulipa gesneriana ), are enzymatically converted to the antimicrobial lactonized aglycons, tulipalins A (PaA) and B (PaB), respectively. We recently identified a PosA‐converting enzyme ( TCEA ) as the first reported member of the lactone‐forming carboxylesterases. Herein, we describe the identification of another lactone‐forming carboxylesterase, PosB‐converting enzyme ( TCEB ), which preferentially reacts with PosB to give PaB. This enzyme was isolated from tulip pollen, which showed high PosB‐converting activity. Purified TCEB exhibited greater activity towards PosB than PosA, which was contrary to that of the TCEA . Novel cDNA ( Tg TCEB 1 ) encoding the TCEB was isolated from tulip pollen. Tg TCEB 1 belonged to the carboxylesterase family and was approximately 50% identical to the Tg TCEA polypeptides. Functional characterization of the recombinant enzyme verified that Tg TCEB 1 catalyzed the conversion of PosB to PaB with an activity comparable with the native TCEB . RT ‐ qPCR analysis of each part of plant revealed that Tg TCEB 1 transcripts were limited almost exclusively to the pollen. Furthermore, the immunostaining of the anther cross‐section using anti‐Tg TCEB 1 polyclonal antibody verified that Tg TCEB 1 was specifically expressed in the pollen grains, but not in the anther cells. N‐terminal transit peptide of Tg TCEB 1 was shown to function as plastid‐targeted signal. Taken together, these results indicate that mature Tg TCEB 1 is specifically localized in plastids of pollen grains. Interestingly, PosB, the substrate of Tg TCEB 1, accumulated on the pollen surface, but not in the intracellular spaces of pollen grains.