Solanum lycopersicum ACC Synthase

Amanda He '16 and Andrew Pearlman '15


I. Introduction

1-aminoacyclopropane 1-carboxylate synthase, also referred to as ACC synthase (ACS), is an enzyme that plays a significant role in synthesizing ethylene, a plant hormone produced in a variety of higher plants [1]. Ethylene is an important hormone for ripening and germination processes.  Ethylene biosynthesis consists of three main steps. The rate-limiting step in ethylene synthesis is the second step, which involves ACS [2]. In the second step, ACS catalyzes the cyclization of S-adenosy-L-methionine (SAM), which produces 5'-methylthioadenosine and ACC [3].
In order for ACS to function, it requires the cofactor, pyridoxal-5'-phosphate (PLP) [4]. PLP has been found to activate catalysis in enzymes, including ACS, by binding to the substrate active site [1].

As the rate-limiting step of ethylene biosynthesis, ACS provides a number of opportunities to inhibit the process. A competitive inhibitor for ACS is aminoethoxyvinylglycine (AVG) [5]. AVG binds to the substrate active site, preventing SAM from interacting with the site [6]. AVG is applied to various fruit-bearing plants to control the rate at which ethylene is produced [7]. Establishing an understanding of how the ACS and the factors it interacts with operate on a molecular level can help identify potential risks of our current agricultural industry. 

II. General Structure

The tomato ACC synthase has two domains, an alpha-beta-alpha sandwich domain and an alpha-beta domain [1]. The is comprised of seven beta strands, which flank nine alpha-helices. The consists of five beta strands and five alpha-helices.

ACS typically appears in a as do most PLP-dependent enzymes. However, there have been no results indicating that monomeric ACS is not catalytically active. So both the monomer and dimer forms are functional. In the dimeric formation, the N-terminal residue in alpha-beta domain makes contact with the . The contact could have a potential role in conformation stabilization or catalysis [1].

Oftentimes, ACS forms complexes with PLP and/or AVG. The ACS-PLP complex uses the dimer form, while ACS-PLP-AVG complex uses the monomer form. When comparing the structural superposition, there was not a significant difference between ACS-PLP and ACS-PLP-AVG, indicating that AVG binding does not significantly change ACS formation [1].

Although it appears more frequently in dimeric form, there is not a substantial difference between the functionality of the monomer and dimer forms because there were not a significant difference in conformational change between the two when the inhibitor, AVG, binds [1].

In ACC synthase, the and the are the main catalytic domains.

III. Cofactor PLP Binding

ACS forms a complex with cofactor, pyridoxal-5'-phosphate , an active form of vitamin B6, in order to be able to catalysis its substrate, S-adenosylmethionine (SAM) [8]. In the dimeric form of ACS, both PLP and SAM are bound to the active site. The PLP ligands interact with the residues of the second ACS monomer, which helps to stabilize the active dimeric form of the enzyme.  PLP forms bonds with ACS with four specific structural components [1]:
Note: Click reset protein between each button for the list below.

  • of the pyridine forms a covalent bond with amine of Lys278
  • of the pyridine forms a H-bond with Asp237
  • of pyridine forms a H-bond with oxygen (O) of Tyr240 and nitrogen (N) of Asn209
  • with Tyr152

Each of these PLP-ACS bonds are a means to stabilizing the structure. For example, in the N1-Asp237 hydrogen bond, the aspartate's negative charge is able to stabilize the protonation state of the pyridine nitrogen[1].

It should be noted that while PLP binding and interactions are largely conserved among most ACC synthase-carrying plant species, the specific amino acid sites where the interactions take place may vary; the information above applies most accurately to tomato ACC synthase.

IV. Substrate SAM Binding

S-adenosylmethionine (SAM) is the substrate that catalyzes the reaction. As substrate binding proceeds, SAM interacts with the PLP-ACS complex [8]. This interaction relies on the participation of a variety of structures and residues. As PLP and SAM both bind to the active sites, proper positioning is crucial. SAM was not crystallized in this pdb file, but all the residues of ACS involved in interactions were highlighted. This is promoted by Ala127, Thr128, Ser275, Ser277, and Arg286, in part due to H-bonding with [9]. There are a number of residue interactions between SAM and ACS that allow for its binding, which include [9]:
  • between the positively-charged sulfonium group within SAM and Glu55
  • from the alpha-carboxylate of SAM to nitrogen (N) of Ala54 and the guanidinium group (g-group) of Arg412
  • between the O2' and O3' of SAM and the guanidinium group (g-group) of Arg157
  • between the adenine ring portion of SAM and a hydrophobic pocket resulting from Pro26, Tyr27, Phe28, and Pro153

V. Inhibitor AVG Binding

Since aminoethoxyvinylglycine (AVG) is a competitive inhibitor of ACC synthase, it is used in the agricultural industry to reduce pre-harvest drop, delay fruit ripening, and reduce ethylene production [6]. As AVG is becoming more commonly used, it is important to identify how AVG interacts with ACC synthase and other parts of the plant.

Note: Click reset complex between each button. 

AVG binds to a location neighboring the cofactor, PLP. The alpha-carboxylate group of AVG forms three hydrogen bonds with PLP at , , and water. Both Ala54 and Arg412 are highly conserved amino acids in ACS because they are also binding sites for SAM. Previous studies identified that substitution at these sites reduces the rate of catalysis. The substitution of the apple ACC synthase equivalent to Arg412 to lysine results in an increase in KM [10]. KM is the measurement of the substrate concentration required for catalysis to occur. The increase in KM when substituting Arg407 for Lys indicates there needs to be a higher concentration of substrate in order for catalysis to occur. As apple and tomato ACC synthase share many similarities, it is likely to assume that the replacement of Arg412 in tomato ACC synthase would have a similar effect.

Additionally, AVG forms a van der Waals contact with . As mentioned earlier, Tyr152 is parallel to the ring of PLP, which allows it to stack, stabilizing the structure.

While the ethylene biosynthesis inhibition effects of AVG has been studied, there is very limited research on how it affects the plant in other manners. A previous study found that AVG presence is correlated to reduced protein synthesis [7]. However, there are not any results correlating AVG use to any consumer harm, so it is continued to be used in fruit production.

VI. References

  1. Huai, Q., Xia, Y., Chen, Y., Callahan, B., Li, N., and Ke, H. 2001. Crystal structures of 1-aminocyclopropane-1-carboxylate (ACC) synthase in complex with aminoethoxyvinylglycine and pyridoxal-5-phosphate provide new insight into catalytic mechanisms. Trends in Plant Science. 10(6): 291-296."

  2. Chae, H.Y. and Kieber, J.J. 2005. Eto Brute? Role of ACS turnover in regulating ethylene biosynthesis. Cell. 11(10): 1383-1394.

  3. Yip, W., Moore, T., and Yang, S.F. 1992. Differential accumulation of transcripts for four tomato 1-aminocyclopropane-1-carboxylate synthase homologs under various conditions. Proc. Natl. Acad. Sci. 89: 2475-2479.

  4. The European Bioinformatics Institute. 2011. "Pyridoxyl-5'-phosphate: PLP Summary." Protein Data Bank in Europe. Web.

  5. Sigma Aldrich. 2014. "L-alpha-[2-(2-aminoethoxy)vinyl]glycine hydrochloride (AVG-HCl)." Web.

  6. Rath, A.C., Kang, I., Park, C., Yoo, W., and Byun, J. 2006. Foliar application of aminoethoxyvinylglycine (AVG) delays fruit ripening and reduces pre-harvest fruit drop and ethylene production of bagged "Kogetsu" apples. Plant Growth Regul. 50: 91-100.

  7. Saltveit, M.E. 2004. Aminoethoxyvinylglycine (AVG) reduces ethylene and protein biosynthesis in excised discs of mature-green tomato pericarp tissue. Postharvest Biology and Technology. 35: 183-190.

  8. Wikipedia, the free encyclopedia. 2014. "1-aminocyclopropane-1-carboxylate synthase." Web.

  9. Jakubowicz, Malgorzata. 2002. Structure, catalytic activity, and evolutionary relationships of 1-aminocyclopropane-1-carboxylate synthase, the key enzyme of ethylene synthesis in higher plants. Acta Biochimica Polonica. 49(3): 757-774.

  10. White, M.F., Vasquez, J., Yang, S.F., and Kirsch, J.F. 1994. Expression of apple 1-aminocyclopropane-1-carboxylate synthase in Escherichia coli: kinetic characterization of wild-type and active-site mutant forms. PNAS. 91(26):12428-12432.

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