Understanding Substrate Specificity of Related Plant Methylesterases(MESs) from Computational Investigations

doi:10.7503/cjcu20150674

Abstract

Abstract:

One of enzyme’s hallmarks is the high specificity to their natural substrates. But our understanding is still lacking concerning how enzymes could achieve high specificity and substrate discrimination. This is the case for a group of related methylesterases(MESs) identified in plants that catalyze reactions of different substrates, including methyl salicylate(MeSA), methyl jasmonate(MeJA) and methyl indole-3-acetate(MeIAA). In this work, a homology model was built for AtMES10(a MeJA esterase),and this model along with the X-ray structure of SABP2(a MeSA esterase) was used to understand their substrate specificity. It is shown that the specificity may be explained based on the simple Lock-and-Key Model(that is, the active site being complementary in shape to the substrate) along with the requirement that the —COO moiety involved in the reaction occupies a position allowing the nucleophilic attack by the catalytic serine(that is, in a reactive configuration). The active site of SABP2 appears not to be complementary in shape to MeJA, and this may lead to a low activity on MeJA. For AtMES10, certain bulky residues in SABP2 are replaced by relatively small residues, allowing the substrate to bind to the active site and to be catalyzed by the enzyme. The results are consistent with the substrate specificity of these two enzymes observed experimentally. Explanations are also provided for the lack of the activities of AtMES10 and SABP2 on MeIAA and the lack of the activity of AtMES10 on MeSA.

Key words: Enzyme catalysis, Substrate specificity, Computer modeling, Protein structure prediction, Methyleste-rase

CLC Number:

O641

TrendMD:

QIAN Ping, ZHAO Nan, CHEN Feng, GUO Hong. Understanding Substrate Specificity of Related Plant Methylesterases(MESs) from Computational Investigations[J]. Chem. J. Chinese Universities, 2015, 36(11): 2283.

Figures/Tables 9

Table 1 Substrate specificities of enzymes*

Name	MelAA	MeSA	MeJA
SABP2	-	+	-
AtMES10	-	-	+

Fig.1 Acylation reaction mechanism of AtMES10 with the catalytic triad involving Ser96, His253 and Asp225 RC:reactant complex; TI: tetrahedral intermediate; AE: acyl-enzyme complex. AtMES10 has esterase activity on methyl jasmonate(MeJA), but no activity was observed on methyl salicylate(MeSA) or methyl indole-3-acetate(MeIAA).

Fig.2 Sequence alignment of SABP2 and AtMES10

Fig.3 Four different conformations of MeJA downloaded from the ZINC database 1: zinc_4654657; 2: zinc_4975331; 3: zinc_4975327; 4: zinc_4975334. The coordinates for the 3D structures can be obtained from ZINC database. The relative stabilities of the conformations are given in Table 2 with 1 and 2 the most and the second most stable conformations, respectively, based on quantum mechanical calculations.

Table 2 Single point energies of four conformers of MeJA in vacuum gas and water solven with the help of polarizable continuum model(PCM) calculated at the level of B3LYP/6-31G(d)//HF/6-31G(d)

Structure code	B3LYP/6-31G(d)//HF/6-31G(d)		PCM
Structure code	SPE/a.u.	ΔE^*/(kJ·mol^-1)	SPE/a.u.	ΔE^*/(kJ·mol^-1)
1	-733.0910853	0	-733.1037684	0
2	-733.0894165	4.39	-733.1024123	3.55
3	-733.086736	11.41	-733.0997755	10.49
4	-733.0857963	13.88	-733.0999091	10.12

Fig.4 Comparison of the homology model of AtMES10(color: orange) on the X-ray structure of SABP2(PDB ID: 1XKL; color: cyan) (A) Superposition of the homology model of AtMES10 on the X-ray structure of SABP2. RMSD=0.062 nm based on the main-chain atoms. SA is shown in space filling; (B) superposition of the catalytic triads for the AtMES10 model and 1XKL. SA is also shown; (C) superposition of some other residues at the binding sites for the AtMES10 model and 1XKL(cyan); (D) X-ray structure of SABP2(1XKL) in space filling. Trp131 and Met149(pink) are located on the surface, SA(grey) can be seen through a channel from the surface to the binding site, see Methods; (E) AtMES10 model in space filling, Trp131 and Met149 are replaced by Ala146 and Val164, respectively.

Fig.5 Docking configuration of 1(the most stable conformer) of MeJA to the active site of SABP2 and AtMES10 model (A) SABP2: bad contacts occur between the MeJA chain and Trp131 or Met149; (B) AtMES10: there are few bad contacts in this case; (C) the active site structure of the AtMES10-MeJA complex after energy minimization. The substrate is well positioned relative to the catalytic triad for the catalysis.

Fig.6 Docking configuration of 2(the second most stable conformer) of MeJA to the active site of SABP2 and AtMES10 model (A) SABP2: the chain of MeJA is located at a different location without severe bad contacts with the residues from the enzyme; (B) AtMES10: without severe bad contacts.

Fig.7 Two docking configurations of indole-3-acetic acid(IAA) to the active site of the AtMES10 model with the carboxylic acid moiety located at the reactive position relative to Ser96

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