Cloning of a cDNA encoding a 3-dehydroquinate synthase from a higher plant, and analysis of the organ-specific and elicitor-induced expression of the corresponding gene

Abstract

cDNA clones for all enzymes of the prechorismate pathway of higher plants have previously been cloned, with the exception of the second enzyme of the pathway, 3-dehydroquinate synthase. Here we describe the isolation of a cDNA encoding a 3-dehydroquinate synthase from tomato which was identified by complementing a 3-dehydroquinate synthase-deficient Escherichia coli strain with a tomato cDNA library. The deduced amino acid sequence contains a putative N-terminal plastid-specific transit peptide, and the sequence of the mature enzyme resembles those of the corresponding bacterial enzymes more than of the fungal enzymes. Sequence identity was even higher between the tomato and E. coli sequences than between the E. coli and other known bacterial sequences. The abundance of 3-dehydroquinate synthase transcripts differ in the organs of tomato plants analyzed. In cultured tomato cells, the abundance of 3-dehydroquinate synthase transcripts increased 9-fold within 4 to 5 h of elicitor treatment.

LeSBT1, a Subtilase from Tomato Plants

“…Reaction products were cloned into Bluescript pSKϪ (Stratagene, La Jolla, CA). A PCR product exhibiting sequence similarity with subtilases was used for the screening of a tomato flower cDNA library in ZAPII (Stratagene) (24). A total of 4 ϫ 10 5 phages were screened on duplicate nitrocellulose filters by hybridization to the radiolabeled cDNA probe (Prime-It system; Stratagene) as described (18).…”

“…In the standard assay, the protease was incubated with a 200-fold molar excess of glucagon and the peptide fragments generated were analyzed by MALDI-TOF mass spectrometry. The 29-amino acid peptide glucagon was preferentially processed carboxyl-terminal of Gln 24 and Gln 20 . After extensive incubation, the resulting peptides (glucagon-(1-24) and glucagon-(1-20)) were further processed carboxyl-terminal of Asp 15 , resulting in glucagon-(1-15) as the final product (Fig.…”

A unique reaction in a common pathway: mechanism and function of chorismate synthase in the shikimate pathway

“…This phenomenon can possibly be explained by a regulation of chorismate synthase activity in planta via light-dependent generation of reduced FMN. However, leucoplasts isolated from roots have also been shown to be capable of synthesizing the three aromatic amino acids (Leuschner and Schultz 1991), and the transcripts of shikimate-pathway enzymes have been shown to be highly abundant in non-photosynthetic tissues (GoÈ rlach et al 1994;Bischo et al 1996). In these tissues, the reducing power cannot be directly derived from photosynthetic electron transport.…”

Isolation of a cDNA from tomato coding for an unregulated, cytosolic chorismate mutase

“…It showed sigmoidal substrate saturation kinetics under appropriate conditions and consisted of at least two identical subunits [24]. In several plant species, the ratio of CM-I to CM-2 type activities has been reported to differ in an organ-and age-specific manner, and only the CM-1 type isozyme was wound-inducible [ 18, 231. While all cDNAs and genes specific for the prechorismate pathway and the tryptophan pathway identified so far encode proteins with N-terminal transit peptides directing the proteins to plastids [4,19,23], the situation for the phenylalanine and tyrosine pathway differs in regard to the presence of two differently compartmentalized CM isozymes. Two CMspecific cDNAs (corresponding to the genes AtCMI and AtCM2) have recently been cloned from Arabidopsis thaliana [6, J.E.…”