Substitutions of Alanine for Cysteine at a Reactive Thiol Site and for Lysine at a Pyridoxal Phosphate Binding Site of 1-Aminocyclopropane-1-carboxylate Deaminase

Murakami, Tetuhide H.; Kiuchi, Miwa; Ito, Hiroyuki; Matsui, Hirokazu; Honma, Mamoru

doi:10.1271/bbb.61.506

Cited by 9 publications

(16 citation statements)

References 6 publications

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“…However, further experiments have shown that substitution of this residue to Ala does not affect the enzymatic activity, suggesting that Cys 165 is not directly involved in the enzymatic activity (42). On the other hand, the replacement of Lys 51 with Ala at the PLP-binding site caused a loss of detectable ACC deamination activity (42). The present structure analysis is consistent with these earlier observations.…”

Section: Resultssupporting

confidence: 90%

“…The enzymatic activity of ACCD is inhibited by modifying Cys 162 of bACCD (which corresponds to Cys 165 of yACCD) with sulfhydryl-modifying reagents (41). However, further experiments have shown that substitution of this residue to Ala does not affect the enzymatic activity, suggesting that Cys 165 is not directly involved in the enzymatic activity (42). On the other hand, the replacement of Lys 51 with Ala at the PLP-binding site caused a loss of detectable ACC deamination activity (42).…”

Section: Resultsmentioning

confidence: 99%

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Crystal Structure of 1-Aminocyclopropane-1-carboxylate Deaminase from Hansenula saturnus

Yao¹,

Ose²,

Sugimoto³

et al. 2000

Journal of Biological Chemistry

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The pyridoxal 5-phosphate (PLP)-dependent enzyme 1-aminocyclopropane-1-carboxylate deaminase (ACCD) catalyzes a reaction that involves a ring opening of cyclopropanoid amino acid, yielding ␣-ketobutyrate and ammonia. Unlike other PLP-dependent enzymes, this enzyme has no ␣-hydrogen atom in the substrate. Thus, a unique mechanism for the bond cleavage is expected. The crystal structure of ACCD from Hansenula saturnus has been determined at 2.0 Å resolution by the multiple wavelength anomalous diffraction method using mercury atoms as anomalous scatterers. The model was built on the electron density map, which was obtained by the density averaging of multiple crystal forms. The final model was refined to an R-factor of 22.5% and an R free -factor of 26.8%. The ACCD folds into two domains, each of which has an open twisted ␣/␤ structure similar to the ␤-subunit of tryptophan synthase. However, in ACCD, unlike in other members of the ␤ family of PLPdependent enzymes, PLP is buried deep in the molecule. The structure provides the first view of the catalytic center of the cyclopropane ring opening.

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Section: Resultssupporting

confidence: 90%

Section: Resultsmentioning

confidence: 99%

Crystal Structure of 1-Aminocyclopropane-1-carboxylate Deaminase from Hansenula saturnus

Yao¹,

Ose²,

Sugimoto³

et al. 2000

Journal of Biological Chemistry

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“…2). K 51 is the PLP-binding site in H. saturnus (Yao et al, 2000) and is required for enzymatic activity in P. fluorescens ACP (Murakami et al, 1997), probably because it functions as the base for proton abstraction (Yao et al, 2000). Structural analysis of the dimeric AcdS protein in H. saturnus (Yao et al, 2000) evidenced a PLP-binding domain (domain I) and an internal, smaller domain (domain II), which were readily identified in the bacterial sequences studied (Fig.…”

Section: Analysis Of Acds Domains and Residues Based On Deduced Protementioning

confidence: 87%

Phylogeny of the 1-aminocyclopropane-1-carboxylic acid deaminase-encoding gene acdS in phytobeneficial and pathogenic Proteobacteria and relation with strain biogeography

Blaha

Prigent‐Combaret

Mirza

et al. 2006

FEMS Microbiology Ecology

250

131

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Deamination of the ethylene precursor 1‐aminocyclopropane‐1‐carboxylic acid (ACC) is a key plant‐beneficial trait found in plant growth‐promoting rhizobacteria (PGPR) and phytosymbiotic bacteria, but the diversity of the corresponding gene (acdS) is poorly documented. Here, acdS sequences were obtained by screening putative ACC deaminase sequences listed in databases, based on phylogenetic properties and key residues. In addition, acdS was sought in 71 proteobacterial strains by PCR amplification and/or hybridization using colony dot blots. The presence of acdS was confirmed in established AcdS+ bacteria and evidenced noticeably in Azospirillum (previously reported as AcdS−), in 10 species of Burkholderia and six Burkholderia cepacia genomovars (which included PGPR, phytopathogens and opportunistic human pathogens), and in five Agrobacterium genomovars. The occurrence of acdS in true and opportunistic pathogens raises new questions concerning their ecology in plant‐associated habitats. Many (but not all) acdS+ bacteria displayed ACC deaminase activity in vitro, including two Burkholderia clinical isolates. Phylogenetic analysis of partial acdS and deduced AcdS sequences evidenced three main phylogenetic clusters, each gathering pathogens and plant‐beneficial strains of contrasting geographic and habitat origins. The acdS phylogenetic tree was only partly congruent with the rrs tree. Two clusters gathered both Betaprotobacteria and Gammaproteobacteria, suggesting extensive horizontal transfers of acdS, noticeably between plant‐associated Proteobacteria.

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“…The existence of the carboxylate group in this position is widely seen in PLP-dependent enzymes with the exception of other members of the TRPS␤ family or alanine racemase from Bacillus stearothermophilus (32 features of PLP enzyme) are expected to be important for the enzymatic activity. A previous paper (33) has also noted that the substitution of alanine for Lys 51 residue at a PLP binding site of pACCD resulted in a complete loss of ACC-deaminating activity and a characteristic absorption spectrum with maxima at 330 and 405 nm and that an addition of ACC to the K51A mutant enzyme caused a decrease of absorbance at 330 nm and an increase at 425 nm, which shifted the maximum from the 405-nm band. This spectral change indicates the formation of an external aldimine between PLP and ACC in the mutant enzyme.…”

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confidence: 81%

Reaction Intermediate Structures of 1-Aminocyclopropane-1-carboxylate Deaminase

Ose

Fujino

Yao

et al. 2003

Journal of Biological Chemistry

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The pyridoxal 5 -phosphate-dependent enzymes have been evolved to catalyze diverse substrates and to cause the reaction to vary. 1-Aminocyclopropane-1-carboxylate deaminase catalyzes the cyclopropane ring-opening reaction followed by deamination specifically. Since it was discovered in 1978, the enzyme has been widely investigated from the mechanistic and physiological viewpoints because the substrate is a precursor of the plant hormone ethylene and the enzymatic reaction includes a cyclopropane ring-opening. We have previously reported the crystal structure of the native enzyme. Here we report the crystal structures of the two reaction intermediates created by the mutagenesis complexed with the substrate. The substrate was validated in the active site of two forms: 1) covalent-bonded external aldimine with the coenzyme in the K51T form and 2) the non-covalent interaction around the coenzyme in the Y295F form. The orientations of the substrate in both structures were quite different form each other. In concert with other site-specific mutation experiments, this experiment revealed the ingenious and unique strategies that are used to achieve the specific activity. The substrate incorporated into the active site is reactivated by a two-phenol charge relay system to lead to the formation of a Schiff base with the coenzyme. The catalytic Lys 51 residue may play a novel role to abstract the methylene proton from the substrate in cooperation with other factors, the carboxylate group of the substrate and the electron-adjusting apparatuses of the coenzyme.The ␣-amino acid with a cyclopropane ring, 1-aminocyclopropane-1-carboxylate (ACC), 1 is known to be a key intermediate in the biosynthesis of the plant hormone ethylene (1), which is responsible for the initiation of fruit-ripening and for regulating many other plant developmental processes. In higher plants, the pathway of ethylene biosynthesis ( Fig. 1) is initiated with the conversion of the methionine to S-adenosylmethionine (2) by methionine adenosyltransferase followed by the conversion of S-adenosylmethionine to ACC by ACC synthase, pyridoxal 5Ј-phosphate (PLP)-dependent enzyme. ACC is oxidized by mononuclear iron dependent-ACC oxydase (3) to ethylene in the final step.In a soil bacterium, Pseudomonas sp. strain ACP, ACC was also converted to ␣-ketobutyrate and ammonia by a bacterial enzyme when ACC was the sole nitrogen source for the growth of this bacterium (4, 5). This degradation is catalyzed by another PLP-dependent enzyme, ACC deaminase (ACCD, EC 4.1.99.4), which has a special ability to break the cyclopropane ring preceding deamination (Fig. 2). The possibility of symbiosis between the plants and the soil bacterium has also been mentioned (6, 7). The introduction of ACCD in higher plants by gene technology reduced the production level of ethylene and delayed the ripening progression of fruits (8, 9). This enzyme has been isolated from a few strains of Pseudomonas species (5, 8, 10, 11), yeast Hansenula saturnus (12), and fungus Penicillium citrinum (13)...

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Substitutions of Alanine for Cysteine at a Reactive Thiol Site and for Lysine at a Pyridoxal Phosphate Binding Site of 1-Aminocyclopropane-1-carboxylate Deaminase

Cited by 9 publications

References 6 publications

Crystal Structure of 1-Aminocyclopropane-1-carboxylate Deaminase from Hansenula saturnus

Crystal Structure of 1-Aminocyclopropane-1-carboxylate Deaminase from Hansenula saturnus

Phylogeny of the 1-aminocyclopropane-1-carboxylic acid deaminase-encoding gene acdS in phytobeneficial and pathogenic Proteobacteria and relation with strain biogeography

Reaction Intermediate Structures of 1-Aminocyclopropane-1-carboxylate Deaminase

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