Crystal structures clarify cofactor binding of plant tyrosine decarboxylase

Biochemical and Biophysical Research Communications xxx (xxxx) xxx

Contents lists available at ScienceDirect

Biochemical and Biophysical Research Communications journal homepage: www.elsevier.com/locate/ybbrc

Crystal structures clarify cofactor binding of plant tyrosine decarboxylase Hang Wang a, Jian Yu a, b, Yasuharu Satoh c, Yusuke Nakagawa a, Ryusuke Tanaka a, Koji Kato a, b, Min Yao a, b, * a b c

Graduate School of Life Science, Hokkaido University, Sapporo, Hokkaido, 060-0810, Japan Faculty of Advanced Life Science, Hokkaido University, Sapporo, Hokkaido, 060-0810, Japan Faculty of Engineering Hokkaido University, Sapporo, 060-8628, Japan

a r t i c l e i n f o

a b s t r a c t

Article history: Received 13 December 2019 Accepted 16 December 2019 Available online xxx

Plant tyrosine decarboxylase (TyrDC) is a group II pyridoxal 50 -phosphate (PLP)-dependent decarboxylase that mainly catalyzes the decarboxylation of tyrosine to tyramine. This is biologically important for diverting essential primary metabolites into secondary metabolic pathways. Intensive studies have characterized the effective of PLP-binding and the substrate specificity of mammalian 3,4dihydroxyphenyl-L-alanine (Dopa) decarboxylases, a member of group II PLP-dependent decarboxylase. However, the characteristics of PLP binding and substrate specificity of plant TyrDCs remain unknown. In this study, we focus on the PLP binding manner, and determined the crystal structures of the apo and PLP binding form of type II TyrDC from Papaver somniferum (PsTyrDCII and PsTyrDCII-PLP). The structures showed that, unlike mammalian Dopa decarboxylase, the binding of PLP does not induce distinct conformational changes of PsTyrDCII regarding the overall structure, but the PLP binding pocket displays conformational changes at Phe124, His203 and Thr262. Combining structural comparation and the obtained biochemical findings, it is demonstrated that PsTyrDCII does not binds PLP tightly. Such characteristics of PLP binding may be required by its catalytic reaction and substrate binding. The activity of TyrDC probably regulated by the concentration of PLP in cells. © 2019 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Keywords: Plant tyrosine decarboxylase Aromatic amino acid decarboxylases PLP-dependent Structure

1. Introduction Many secondary metabolites in higher plants need to utilize aromatic amino acids as precursors [1]. The enzymes responsible for diverting these essential primary metabolites into secondary metabolic pathways are thus biologically important. In various organisms, aromatic amino acid decarboxylases (AADCs) are one type of enzyme associated with amino acid metabolism. Mammalian and insect AADCs catalyze the decarboxylation of 3,4-dihydroxyphenyl-L-alanine (Dopa) (named Dopa decarboxylases) and 5-hydroxy-L-tryptophan (5-HTP) (named 5-HTP decarboxylases) to produce dopamine and 5-hydroxytryptamine (serotonin) which serve as key neurotransmitters [2]. In plants, AADCs are involved in the biosynthesis of several different secondary metabolites. Among these, tyrosine decarboxylase (TyrDC),

* Corresponding author. Graduate School of Life Science, Hokkaido University, Sapporo, Hokkaido, 060-0810, Japan. E-mail address: [email protected] (M. Yao).

which is responsible for synthesizing tyramine from tyrosine, has been implicated in the biosynthesis of plant benzylisoquinoline alkaloids (BIAs) [3]. In the initial step of BIA biosynthesis, the central intermediate, (S)-norcoclaurine, is generated by the condensation of dopamine and 4-hydroxyphenylacetaldehyde (4HPAA). Both dopamine and 4-HPAA are derived from tyrosine by TyrDC because either the decarboxylation of Dopa or the hydroxylation of tyramine results in dopamine synthesis. The 4-HPAA would also be produced in a similar manner through the decarboxylation of 4-hydroxyphenylpyruvate or the oxidation of tyramine [4]. Furthermore, tyramine can also contribute to medicine as a significant intermediate of bezafibrate [5]. Therefore, TyrDC has attracted attention as an enzyme for producing tyramine. On the basis of the sequence similarity and domain conservation, nine different pyridoxal 50 -phosphate (PLP)-dependent AADCs have been classified into four groups. TyrDC belongs to group II decarboxylase, which includes L-glutamate decarboxylase, L-histidine decarboxylase, L-tryptophan decarboxylase, and Dopa decarboxylases [6]. Plant and mammalian AADCs share amino acid

https://doi.org/10.1016/j.bbrc.2019.12.077 0006-291X/© 2019 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Please cite this article as: H. Wang et al., Crystal structures clarify cofactor binding of plant tyrosine decarboxylase, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.12.077

2

H. Wang et al. / Biochemical and Biophysical Research Communications xxx (xxxx) xxx

sequence identity and display remarkable similarities in domain structure, molecular mass, and kinetic properties [3]. However, in terms of substrate specificity, these enzymes show distinct differences. Mammalian AADCs are highly specific to Dopa and 5-HTP, but do not react with L-tyrosine (Tyr), L-phenylalanine (Phe), and Ltryptophan (Trp). In contrast, Tyr/Dopa, Trp, and Phe can be specifically distinguished individually by different plant AADCs with different reactional activity [7]. For example, AADC from the plant Catharanthus roseus specifically reacts with Trp [8], whereas that from Petunia hybrida only recognizes Phe as a substrate [9]. To date, mammalian AADCs, Dopa decarboxylases (DopaDCs), have attracted substantial attention. For example, the structures of pig kidney DopaDC (PkDopaDC) and human DopaDC (HuDopaDC) have been studied in detail. The apo and PLP binding form of mammalian DopaDCs show strikingly different conformations [10]. However, the characteristics of PLP-binding and specific substrate binding of plant TyrDC remain unknown. In this study, we present the crystal structures of type II TyrDC from Papaver somniferum (PsTyrDCII), in apo and PLP binding form (PsTyrDCII-PLP). Unlike mammalian DopaDCs, the structures of PsTyrDCII do not show distinct conformational changes between the apo and PLP binding form. Furthermore, the purified recombinant PsTyrDCII showed almost no activity when no cofactor PLP was added. Taken structural information and biochemical findings together, our results indicate that PsTyrDCII binds weakly to cofactor PLP, which may be necessary for its catalytic reaction and substrate binding. 2. Materials and methods

sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDSPAGE) (Fig. 1A). These proteins were concentrated using Amicon Ultra-15 centrifugal filters (Millipore), and the protein concentration was determined using a Nanodrop 2000 spectrophotometer (Thermo Scientific). Finally, the purified proteins were stored at 80  C.

2.3. Crystallization The purified PsTyrDCII proteins were concentrated to 10 mg mL1 for crystallization. Initial screening was performed by the sitting-drop vapor-diffusion method against 960 (96  10 kits) conditions using commercial kits at 20  C. Drops consisting of 0.75 mL of protein and an equal volume of reservoir buffer were equilibrated against 75 mL of reservoir solution. The initial crystal appeared in conditions featuring 1.1 M sodium malonate, 0.1 M HEPES pH 7.0, and 0.5% (v/v) Jeffamine ED-2001. To improve the crystal quality, additional experiments were carried out by mixing 2 mL of protein solution with 2 mL of reservoir solution [0.9 M sodium malonate, 0.1 M HEPES pH 7.0, 0.5% (v/v) Jeffamine ED-2001, and 0.01 M TCEP-HCl] and equilibrating against 500 mL of reservoir solution using the hanging-drop vapor-diffusion method (Fig. 1B). To obtain the PsTyrDCII-PLP crystal, the PsTyrDCII crystal prepared by the above method was soaked in buffer containing different concentrations of PLP. Finally, the PsTyrDCII-PLP crystal was obtained under conditions of 0.9 M disodium malonate, 0.1 M HEPES pH 8.0, 0.01 M TCEP-HCl, 2% glycerol, and 9 nM PLP for soaking 18 h at 20  C.

2.1. Chemical compounds, screening kits, and bacterial strains Tyrosine, tyramine, and PLP were purchased from SigmaAldrich Co. LLC (St. Louis, MO, USA). Crystallization screening kits were purchased from Hampton Research (Aliso Viejo, CA, USA). The bacterial strains used in this study were Escherichia coli XL10-gold and BL21 (DE3) (Novagen). 2.2. Expression and purification of PsTyrDCII The gene fragment encoding PsTyrDCII was amplified using high-fidelity DNA polymerase KOD Plus Neo (Toyobo). The primers 50 -GGGAATTCCATATGGGCTCTCTGAATACGGAAG-30 and 50 CCCAAGCTTTTAACTACTGAAGTCCGCCTCAC-30 were designed to contain NdeI and HindIII sites (underlined), respectively. The amplified fragment was inserted into the corresponding site of pET28b (Novagen) to generate an N-terminal 6histidine-tagged protein. Recombinant E. coli BL21 (DE3) harboring pET28b-PsTyrDCII was cultivated in Luria Broth (LB) medium containing 100 mg L1 kanamycin at 37  C until the optical density at 600 nm (OD600) reached about 0.7. Isopropyl-D-1-thiogalactopyranoside was then added to a final concentration of 0.5 mM and further cultured at 18  C to induce protein expression. After induction for 15e16 h, the cells were harvested by centrifugation at 4500  g for 20 min at 4  C and resuspended in buffer A (50 mM TriseHCl pH 8.0, 500 mM NaCl). Cell extracts were obtained by sonication followed by centrifugation at 25,000  g for 30 min at 4  C to remove cell debris. The resulting supernatant was loaded onto a HisTrap HP column (GE Healthcare, USA), which was pre-equilibrated with buffer A. Subsequently, the retained protein was eluted with a 20e500 mM imidazole gradient in buffer B (50 mM TriseHCl pH 8.0, 500 mM NaCl, and 500 mM imidazole). Furthermore, proteins were dialyzed and then purified by Superdex 200 16/60 column chromatography (GE Healthcare, USA) in a buffer consisting of 50 mM TriseHCl pH 8.0 and 100 mM NaCl for enzymatic activity assay or in a buffer A for crystallization. The purified proteins were confirmed by 15%

2.4. Data collection, structure determination, and refinement For X-ray diffraction experiments, the crystals were soaked in a cryoprotectant solution of reservoir solution with a final concentration of 20% glycerol for several seconds. The crystals were mounted in a cryo-loop and flash-frozen under a stream of gaseous nitrogen. Diffraction data sets were collected on beamline BL44XU at SPring-8 (Harima, Japan) for PsTyrDCII and BL-1A at Photon Factory (Tsukuba, Japan) for PsTyrDCII-PLP. For PsTyrDCII, 360 images were collected with an oscillation angle of 0.5 and an X-ray exposure time of 1 s. For PsTyrDCII-PLP, 1800 images were collected with an oscillation angle of 0.1 and an X-ray exposure time of 1 s. All data sets were indexed and integrated using XDS [11], and scaled and merged using AIMLESS [12] from the CCP4 program suite [13]. The structure of PsTyrDCII was determined by molecular replacement (MR) with Phaser [14] of Phenix program suite [15]. As the search model, the PkDopaDC structure (Protein Data Bank entry number, PDB ID:1JS3) [16] was used, which shares sequence identity of 44% with PsTyrDCII. The model of PsTyrDCII was rebuilt using phenix.autobuild [17] of the Phenix program suite [18]. Structural refinement was performed using phenix.refine [17] of the Phenix program suite, following manual building and modification with Coot [19]. The structure of PsTyrDCII-PLP was determined by rigid-body refinement using coordinates of PsTyrDCII with phenix.refine; density maps of both Fo-Fc and 2Fo-Fc showed a blocklike PLP shape at the active site. After several rounds of structural refinement, PLP molecules were built and structural refinement of PsTyrDCII-PLP was performed like that of PsTyrDCII described above. Data collection and refinement statistics are summarized in Table 1. Figures of structures were generated using PyMOL [20]. The refined atomic coordinates and experimental structural factors of PsTyrDCII and PsTyrDCII-PLP have been deposited in the PDB with entry numbers of 6LIU and 6LIV, respectively.

Please cite this article as: H. Wang et al., Crystal structures clarify cofactor binding of plant tyrosine decarboxylase, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.12.077

H. Wang et al. / Biochemical and Biophysical Research Communications xxx (xxxx) xxx

3

Fig. 1. Purification and crystallization of PsTyrDCII. (A) The peak identified by size exclusion chromatography of PsTyrDCII was confirmed by SDS-PAGE. (B) Image of PsTyrDCII crystals.

Table 1 X-ray data collection and structure-refinement statistics.

PDB ID Data collection Wavelength Resolution range Space group Unit cell parameters

PsTDCII

PsTDCII-PLP

6LIU

6LIV

0.90000 50e2.798 P21212 217.159 118.043 180.864 90 90 90 7.40 (7.60) 99.9 (99.4) 7.37 (2.07) 52.31 22.1 (98.0) 99.2 (81.6)

1.10000 50e2.312 P21212 218.057 117.911 180.414 90 90 90

Redundancy 6.96 (6.66) Completeness (%) 99.68 (97.81) Mean I/sigma(I) 7.12 (1.70) Wilson B-factor 32.07 a R-meas 21.4 (88.7) CC1/2 99.1 (72.1) Refinement Rwork/Rfree (%)b 24.02/27.88 23.29/25.59 No. non-H atoms 22059 23784 Proteins 21935 22766 Ligand e 198 water 124 1018 RMS (bonds) 0.006 0.005 RMS (angles) 0.72 0.673 Ramachandran favoured (%) 96.96 97.07 allowed (%) 3.04 2.86 Average B-factor 56.63 45.32 Proteins 56.65 45.73 Ligand e 36.93 water 54.50 37.36 P P P P a . Rmeas ¼ hkl{N(hkl)/[N(hkl)-1]}1/2 i|Ii(hkl)-|/ hkl iIi(hkl), where and N(hkl) are the mean intensity of a set of equivalent reflections and the multiplicity, respectively. The highest resolution shell is shown in parentheses. P P b . Rwork ¼ hkl||Fobs|-|Fcalc||/ hkl|Fobs|, Rfree was calculated for 5% randomly selected test sets that were not used in the refinement.

2.5. Enzymatic activity assay The reaction assay was performed using a mixed (1 mL) reaction solution containing 50 mM TriseHCl pH 8.0, and 0.5 mM tyrosine with/without 0.1 mM PLP. The mixture was first pre-incubated for 2 min at 35  C; then, 0.675 mM purified recombinant PsTyrDCII was added and the reaction proceeded. The reaction was carried out for 10 min at 35  C and stopped by adding 100% methanol. Reaction products (10 mL) were analyzed using the HPLC system JASCO LC-4000 (JASCO, Tokyo, Japan) equipped with a Discovery HS F5 column (150 mm length  2.1 mm internal diameter, 3 mm; Sigma-Aldrich Co. LLC, St. Louis, MO, USA). Buffer C [MilliQ

containing 0.1% (v/v) formic acid] and Buffer D [acetonitrile containing 0.1% (v/v) formic acid] were used as the mobile phase. The samples were eluted at 35  C and a flow rate of 0.2 mL min1 with an increasing concentration of buffer D as follows: 0% e 50%, 0 e 25 min. 3. Results and discussion 3.1. Overall structure of PsTyrDCII The structure of PsTyrDCII was determined by an MR method at 2.798 Å resolution, and the final structure was refined to Rfree/Rwork

Please cite this article as: H. Wang et al., Crystal structures clarify cofactor binding of plant tyrosine decarboxylase, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.12.077

4

H. Wang et al. / Biochemical and Biophysical Research Communications xxx (xxxx) xxx

of 24.02/27.88%. Like other DopaDCs and TyrDCs, PsTyrDCII forms a homodimer with two active sites each contributed to by two monomers (Fig. 2A), and three homodimers (molAB, molCD, and molEF) exist in an asymmetric unit. Each monomer was built, except the N-terminal residues (16 residues in MolA, B, and C; 15 residues in MolD; 23 residues in MolE; 18 residues in MolF), a loop (Ala353 e Gln358 in MolA and C; Asn351 e Gln358 in other monomers), and a fragment (Lys433 e Asn456 in all monomers). Hence, we use dimer molAB for the following explanation and discussion. Structural comparison of PsTyrDCII with other enzymes using the Dali program [21] showed that PsTyrDCII dimer shares its overall structure with other known structures of AADCs, with root mean square difference (RMSD) values of 0.359 Å for 826 Ca-atoms of PsTyrDCI-PLP complexed with Tyr (PDBID: 6EEM; 81% sequence identity) [22], 0.874 Å for 780 Ca-atoms of PkDopaDC-PLP (PDBID: 1JS6; 44% sequence identity) [16], 2.839 Å for 703 Ca-atoms of HuDopaDC (PDBID: 3RBL; 42% sequence identity) [10], 2.189 Å for 797 Ca-atoms of drDopaDC-PLP (PDBID: 3K40; 41% sequence identity) [23], and 11.797 Å for 637 Ca-atoms of LbTyrDC (PDBID: 5HSI; 18% sequence identity) [24]. Those structures also form a similar active pocket with a slightly different conformation. Previous studies reported that an important long and flexible loop (Ser343eAsp361) named catalytic loop including one crucial catalytic residue Tyr348 around the active site was partially visualized only in PsTyrDCI-PLP [22]. In our apo form structure, the catalytic loop was partly visualized (Ser343eArg350) with a different conformation, indicating its flexibility. In addition, compared with the results of Torrens et al. the catalytic Tyr348 in our apo form was far away from the active pocket.

3.2. Characterization of PLP binding of PsTyrDCII Using recombinant PsTyrDCII, we could not obtain the crystal structure of PsTyrDCII complexed with cofactor PLP without the addition of PLP reagent during purification and crystallization, suggesting that PsTyrDCII may bind weakly to PLP and require a high concentration of PLP for its activity. Therefore, we performed enzymatic activity assay, with the results showing that, without

adding extra PLP, purified PsTyrDCII displayed extremely low activity (3.5% ± 0.7%) (Fig. 3), indicating that almost no purified recombinant PsTyrDCII captured PLP during overexpression. To obtain the structure of PsTyrDCII-PLP, we soaked the crystals in a high concentration of PLP (molar ratio of PsTyrDCII:PLP ¼ 1:25), and consequently obtained the structure of PsTyrDCII complexed with PLP. Such PLP concentration requirement was also obtained for the mammalian AADC HuDopaDC [10]. Similar to the PsTyrDCII structure, each monomer of three PsTyrDCII-PLP dimers (molAB, molCD, and molEF) was built, except the N-terminal residues, a loop and a fragment explained above, which was confirmed by omit-map. In the structure of PsTyrDCIIPLP, the PLP molecule was located on each monomer and covalently bound to the ε-amino group of Lys319 to form an internal aldimine, lysine-pyridoxal-5-phosphate (LLP), via a Schiffebase interaction (Fig. 2B). The PLP molecule was also coordinated by hydrogen bonds with conserved Asp287, almost conserved Asn316, and unconserved Ser370 of the other monomer (Ser370MolB). By comparing the structures of PsTyrDCII-PLP and PsTyrDCII, it was clear that binding of PLP does not induce distinct conformational changes in the overall structure (RMSD value of 0.24 Å for 878 Ca atoms), whereas mammalian DopaDC enzymes have a bivalve conformation for which the binding of PLP induces a change of the open state to a closed one. Although the overall structures of PsTyrDCII-PLP and PsTyrDCII are very similar, conformational changes were found at the active site. In the PLP binding pocket, the side chain of conserved Thr262 turned to make space for PLP binding (Fig. 4A). The side chain of Phe124 of the other monomer (Phe124MolB), which is conserved in TyrDCs and DopaDCs forms different conformations in different stages of the TyrDC and DopaDC structures. In our structures, before binding to PLP, the phenyl ring of Phe124MolB was positioned at the opposite site of the phosphate group of PLP, whereas the phenyl ring of Phe124MolB turned close to PLP in PsTyrDCII-PLP. In the structures of PsTyrDCIPLP and PkDopaDC-PLP (Phe121MolB in PsTyrDCI, Phe103MolB in PkDopaDC) complexed with substrate/inhibitor, the phenyl ring of Phe124MolB turned toward the substrate/inhibitor to undergo static interaction. Owing to the movement of Phe124MolB, the entrance of

Fig. 2. Overall crystal structures of PsTyrDCII and PsTyrDCII-PLP. (A) The dimer structure of PsTyrDCII is shown as a ribbon model. Two monomers were colored in yelloworange and cyan. Close-up view is an active site, and the catalytic loop Ser343molB - Arg350molB (marine) is marked. (B) A ribbon representation of the PsTyrDCII-PLP dimer (pink and limon). PLP covalently bound to Lys319 (LLP) is shown in stick with Fo-Fc map contoured at the 3s level (light-blue) in the closed-up view. The dashed lines show hydrogen binding of PLP with labeled residues.

Please cite this article as: H. Wang et al., Crystal structures clarify cofactor binding of plant tyrosine decarboxylase, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.12.077

H. Wang et al. / Biochemical and Biophysical Research Communications xxx (xxxx) xxx

5

Fig. 3. The results of recombinant PsTyrDCII enzymatic activity assay by HPLC analysis. (A) The plots of HPLC analysis. The reaction product from tyrosine harboring recombinant PsTyrDCII without (a) and with (b) additional PLP. (c) and (d) are references which only injected the compounds of tyramine and tyrosine, respectively. (B) Relative activity of recombinant PsTyrDCII with additional PLP or without additional PLP.

Fig. 4. Structural comparison of PsTyrDCII and PsTyrDCII-PLP with other AADCs. (A) Superposition of active site of PsTyrDCII and PsTyrDCII-PLP dimer. PsTyrDCII and PsTyrDCII-PLP dimer are colored in the same as in Fig. 2. LLP (gray for C atoms), the side chains of Phe124molB, Thr262 and His203 are showed as sticks. In stick presentation, C atoms of Phe124molB, His203, and Thr262 of PsTyrDCII and PsTyrDCII-PLP are colored in the same as in Fig. 2, the O and N atoms are colored in red and blue, respectively. (B) Comparison with other AADCs. The active site of PsTyrDCII-PLP dimer is showed in the same way as in (A). The loop W365molB e R374molB of PsTyrDCII-PLP is superposed with that of PsTyrDCI-PLP (lightorange), CrTrpDC-PLP (yellow) and PkDopaDC-PLP (violetpurple). The side-chain of central residues Ala368molB e Ser370molB of PsTyrDCII-PLP and their related residues in PsTyrDCI/CrTrpDC/PkDopaDC are shown as sticks and colored in the same way as in (A). The sequence alignment of the loop (Trp365 e Arg374) among six enzymes of AADCs (members of group II pyridoxal 50 -phosphate (PLP)-dependent decarboxylase). PsTyrDCI is type I tyrosine decarboxylase from Papaver somniferum. CrTrpDC is tryptophan decarboxylase from Catharanthus roseus. PhPAAS is phenylacetaldehyde synthase from Petunia hybrida. HuDopaDC and PkDopaDC are Dopa decarboxylases from human and pig kidney, respectively. The number of residues is referenced to PsTyrDCII.

the active site in PsTyrDCII-PLP became narrower, compared with that in PsTyrDCII (Fig. 4A). Conformational change between PsTyrDCII and PsTyrDCII-PLP was also identified in a conserved loop (Ser198 e His203) on top of the pyridoxal group, especially for His203. Since the imidazole ring of His203 formed a pi-stacking interaction with the phenyl ring of PLP, there was almost 90 rotation of the His203 imidazole ring between PsTyrDCII and PsTyrDCII-PLP (Fig. 4A). Such changes of the three residues suggest

that Phe124MolB, His203, Thr262 play important roles in binding and stabilizing cofactor PLP. Interestingly, the center (Ala368MolB e Ser370MolB) of a loop (Trp365MolB e Arg374MolB) that interacts with the phosphate group of PLP to anchor PLP did not show conformational changes between PsTyrDCII and PsTyrDCII-PLP (Fig. 4B) but differed from the structures of other DopaDCs and TyrDCs complexed with inhibitor/ substrate. Although this central part shows sequence variation, the

Please cite this article as: H. Wang et al., Crystal structures clarify cofactor binding of plant tyrosine decarboxylase, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.12.077

6

H. Wang et al. / Biochemical and Biophysical Research Communications xxx (xxxx) xxx

residues at both sides of this loop are conserved. Such structural and sequence features may correspond to each enzyme’s manner of PLP binding to ensure substrate specificity. Combining our structures together with other TyrDC/DopaDC structural and biochemical information, it is demonstrated that, unlike other PLP-dependent enzymes, PLP binding of PsTyrDCII is relatively flexible and weak. This may be necessary for the catalytic reaction and substrate binding. The activity of TyrDC can be regulated by the concentration of PLP in the cell. Declaration of competing interest

[8]

[9]

[10]

The authors declare no conflict interest. Acknowledgments

[11] [12] [13]

We are grateful for synchrotron beam time at Photon Factory and SPring-8 and for help from the beamline staff. We are also grateful to Dr. Taek Soon Lee (Joint BioEnergy Institute, USA) for providing PsTyrDCII expression plasmid. This work was supported by grants from the Japanese Ministry of Education, Culture, Sports, Science and Technology (MEXT) and the Japan Society for the Promotion of Science Grants-in-Aid for Scientific Research on Innovative Areas (No. 17H05424 and 19H04633 (to M.Y.)). H. Wang was supported by the Ministry of Education, Culture, Sports, Science and Technology of Japan and by the China Scholarship Council. Appendix A. Supplementary data

[14]

[15]

[16]

Supplementary data to this article can be found online at https://doi.org/10.1016/j.bbrc.2019.12.077.

[17]

References

[18]

[1] K.M. Oksman-Caldentey, D. Inze, Plant cell factories in the post-genomic era: new ways to produce designer secondary metabolites, Trends Plant Sci. 9 (2004) 433e440. [2] R.B. Hodgetts, S.L. O’Keefe, Dopa decarboxylase: a model gene-enzyme system for studying development, behavior, and systematics, Annu. Rev. Entomol., pp. 259-284. [3] P.J. Facchini, K.L. Huber-Allanach, L.W. Tari, Plant aromatic L-amino acid decarboxylases: evolution, biochemistry, regulation, and metabolic engineering applications, Phytochemistry 54 (2000) 121e138. [4] P.J. Facchini, V. De Luca, Phloem-specific expression of tyrosine/dopa decarboxylase genes and the biosynthesis of isoquinoline alkaloids in opium poppy, Plant Cell 7 (1995) 1811e1821. [5] T. NishimakiMogami, K. Suzuki, E. Okochi, A. Takahashi, Bezafibrate and clofibric acid are novel inhibitors of phosphatidylcholine synthesis via the methylation of phosphatidylethanolamine, Biochim. Biophys. Acta Lipids Lipid Metab. 1304 (1996) 11e20. [6] E. Sandmeier, T.I. Hale, P. Christen, Multiple evolutionary origin OF PYRIDOXAL-5’-PHOSPHATE-DEPENDENT amino-acid decarboxylases, Eur. J. Biochem. 221 (1994) 997e1002. [7] T. Koyanagi, A. Nakagawa, H. Sakurama, K. Yamamoto, N. Sakurai, Y. Takagi,

[19] [20] [21] [22]

[23]

[24]

H. Minami, T. Katayama, H. Kumagai, Eukaryotic-type aromatic amino acid decarboxylase from the root colonizer Pseudomonas putida is highly specific for 3,4-dihydroxyphenyl-L-alanine, an allelochemical in the rhizosphere, Microbiology-Sgm 158 (2012) 2965e2974. M.P. Torrens-Spence, M. Lazear, R. von Guggenberg, H.Z. Ding, J.Y. Li, Investigation of a substrate-specifying residue within Papaver somniferum and Catharanthus roseus aromatic amino acid decarboxylases, Phytochemistry 106 (2014) 37e43. Y. Kaminaga, J. Schnepp, G. Peel, C.M. Kish, G. Ben-Nissan, D. Weiss, I. Orlova, O. Lavie, D. Rhodes, K. Wood, D.M. Porterfield, A.J.L. Cooper, J.V. Schloss, E. Pichersky, A. Vainstein, N. Dudareva, Plant phenylacetaldehyde synthase is a bifunctional homotetrameric enzyme that catalyzes phenylalanine decarboxylation and oxidation, J. Biol. Chem. 281 (2006) 23357e23366. G. Giardina, R. Montioli, S. Gianni, B. Cellini, A. Paiardini, C.B. Voltattorni, F. Cutruzzola, Open conformation of human DOPA decarboxylase reveals the mechanism of PLP addition to Group II decarboxylases, Proc. Natl. Acad. Sci. U. S. A 108 (2011) 20514e20519. W. Kabsch, Xds, Acta Crystallogr D Biol Crystallogr 66 (2010) 125e132. P.R. Evans, G.N. Murshudov, How good are my data and what is the resolution? Acta Crystallogr D Biol Crystallogr 69 (2013) 1204e1214. M.D. Winn, C.C. Ballard, K.D. Cowtan, E.J. Dodson, P. Emsley, P.R. Evans, R.M. Keegan, E.B. Krissinel, A.G.W. Leslie, A. McCoy, S.J. McNicholas, G.N. Murshudov, N.S. Pannu, E.A. Potterton, H.R. Powell, R.J. Read, A. Vagin, K.S. Wilson, Overview of the CCP4 suite and current developments, Acta Crystallogr. Sect. D Biol. 67 (2011) 235e242. A.J. McCoy, R.W. Grosse-Kunstleve, P.D. Adams, M.D. Winn, L.C. Storoni, R.J. Read, Phaser crystallographic software, J. Appl. Crystallogr. 40 (2007) 658e674. D. Liebschner, P.V. Afonine, M.L. Baker, G. Bunkoczi, V.B. Chen, T.I. Croll, B. Hintze, L.W. Hung, S. Jain, A.J. McCoy, N.W. Moriarty, R.D. Oeffner, B.K. Poon, M.G. Prisant, R.J. Read, J.S. Richardson, D.C. Richardson, M.D. Sammito, O.V. Sobolev, D.H. Stockwell, T.C. Terwilliger, A.G. Urzhumtsev, L.L. Videau, C.J. Williams, P.D. Adams, Macromolecular structure determination using Xrays, neutrons and electrons: recent developments in Phenix, Acta Crystallogr. Sect. D Biol. 75 (2019) 861e877. P. Burkhard, P. Dominici, C. Borri-Voltattorni, J.N. Jansonius, V.N. Malashkevich, Structural insight into Parkinson’s disease treatment from drug-inhibited DOPA decarboxylase, Nat. Struct. Biol. 8 (2001) 963e967. T.C. Terwilliger, R.W. Grosse-Kunstleve, P.V. Afonine, N.W. Moriarty, P.H. Zwart, L.W. Hung, R.J. Read, P.D. Adams, Iterative model building, structure refinement and density modification with the PHENIX AutoBuild wizard, Acta Crystallogr. Sect. D Biol. Crystallogr. 64 (2008) 61e69. P.D. Adams, P.V. Afonine, G. Bunkoczi, V.B. Chen, I.W. Davis, N. Echols, J.J. Headd, L.W. Hung, G.J. Kapral, R.W. Grosse-Kunstleve, A.J. McCoy, N.W. Moriarty, R. Oeffner, R.J. Read, D.C. Richardson, J.S. Richardson, T.C. Terwilliger, P.H. Zwart, PHENIX: a comprehensive Python-based system for macromolecular structure solution, Acta Crystallogr D Biol Crystallogr 66 (2010) 213e221. P. Emsley, K. Cowtan, Coot: model-building tools for molecular graphics, Acta Crystallogr D Biol Crystallogr 60 (2004) 2126e2132. W.L. Delano, The PyMOL molecular graphics system. http://www.pymol.org, 2002. L. Holm, P. Rosenstrom, Dali server: conservation mapping in 3D, Nucleic Acids Res. 38 (2010) W545eW549. M.P. Torrens-Spence, Y.-C. Chiang, T. Smith, M.A. Vicent, Y. Wang, J.-K. Weng, Structural Basis for Independent Origins of New Catalytic Machineries in Plant AAAD Proteins, 2018. Q. Han, H. Ding, H. Robinson, B.M. Christensen, J. Li, Crystal structure and substrate specificity of Drosophila 3,4-dihydroxyphenylalanine decarboxylase, PLoS One 5 (2010) e8826. H. Zhu, G. Xu, K. Zhang, X. Kong, R. Han, J. Zhou, Y. Ni, Crystal structure of tyrosine decarboxylase and identification of key residues involved in conformational swing and substrate binding, Sci. Rep. 6 (2016) 27779.

Please cite this article as: H. Wang et al., Crystal structures clarify cofactor binding of plant tyrosine decarboxylase, Biochemical and Biophysical Research Communications, https://doi.org/10.1016/j.bbrc.2019.12.077