Fig. S1 A622 1 MVVSEKSKILIIGGTGYIGKYLVETSAKSGHPTFALIRESTLKNPEKSKLIDTFKSYGVT 60 A622L 1..................................V.......V................. 60 A622.

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Presentation transcript:

Fig. S1 A622 1 MVVSEKSKILIIGGTGYIGKYLVETSAKSGHPTFALIRESTLKNPEKSKLIDTFKSYGVT 60 A622L V V A LLFGDISNQESLLKAIKQVDVVISTVGGQQFTDQVNIIKAIKEAGNIKRFLPSEFGFDVD 120 A622L A A HARAIEPAASLFALKVRIRRMIEAEGIPYTYVICNWFADFFLPNLGQLEAKTPPRDKVVI 180 A622L 121..H K A FGDGNPKAIYVKEEDIATYTIEAVDDPRTLNKTLHMRPPANILSFNEIVSLWEDKIGKTL 240 A622L MK E A EKLYLSEEDILQIVQEGPLPLRTNLAICHSVFVNGDSANFEVQPPTGVEATELYPKVKYT 300 A622L H......M...V I..S A TVDEFYNKFV 311 A622L Y (Identity=95.5%, Similarity=98.4%)

Fig. S A622 A622L (73.0%)(85.9%)(68.4%)(43.3%) (98.4%) (98.5%)(97.1%)(96.6%) (93.4%) (nt)

Fig. S3 Red: A622 Blue: ObEGS1+NADPH Green: PtPCBER

Supplementary Figures Figure S1. Alignment of the amino acid sequences of A622 and A622L. Dots indicate the A622L residues that are identical to those of the A622 sequence. Figure S2. Diagram of the A622 and A622L genes. Filled boxes indicate coding exons, whereas the lines show introns. The numbers indicate the lengths of the exons and introns in nucleotides. The sequence identity values between corresponding gene segments are indicated in parentheses. The A622L genomic sequence was deposited in GenBank under the accession number AB Figure S3. Superposition of the polypeptide-chain backbones of A622, ObEGS1 (PDB entry no. 2R6J), and PtPCBER (PDB entry no. 1qyc). The color coding is shown in the inset. The NADPH cofactor in ObEGS1 is shown in the ball mode. The structural modeling of A622 protein and the comparison with other PIP-family proteins were carried out using MODELLER 8v2 (University of California San Francisco, San Francisco, CA) and PyMol version 0.99rc6 (DeLano Scientific LLC, Palo Alto, CA) software, respectively. Supplementary Materials and Methods Determination of the A622L genomic and cDNA sequences Since the A622L genomic sequence in the N. tabacum genome database ( was incomplete, genomic PCR was carried out with the primers 5’- CCTCCACCTTAACCCGAAGC and 5’-TGCAGATTGATGTCGACAAC, using 10 ng of N. tabacum genomic DNA and TaKaRa Ex Taq DNA polymerase (TaKaRa Bio) under the following conditions: 30 cycles of 94 °C for 30 sec, 55 °C for 30 sec, and 72 °C for 1.5 min. The PCR product was sequenced directly by using 5’-TCAGAGAAAGCACACTCGT and 5'-GCATATGGCCAAATTGACT. To determine the cDNA sequence of A622L, RT-PCR was carried out with 5’- CCTCCACCTTAACCCGAAGC and 5’-TGCAGATTGATGTCGACAAC, using 1 ng of N. tabacum root tissue cDNA and TaKaRa Ex Taq DNA polymerase (TaKaRa Bio) under the following conditions: 30 cycles of 94 °C for 30 sec, 55 °C for 30 sec, and 72 °C for 1 min. The PCR product was cloned into pGEM-T easy vector (Promega, Madison, WI), and sequenced using 5’-TCAGAGAAAGCACACTCGT and 5'- GCATATGGCCAAATTGACT.