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Identification and Characterization of Four Missense Mutations in Brown midrib 12 (Bmr12), the Caffeic O-Methyltranferase (COMT) of Sorghum

Sattler, Scott E., Palmer, Nathan A., Saballos, Ana, Greene, Ann M., Xin, Zhanguo, Sarath, Gautam, Vermerris, Wilfred, Pedersen, Jeffrey F.
Bioenergy research 2012 v.5 no.4 pp. 855
Sorghum bicolor, amino acids, biofuels, biosynthesis, energy crops, enzyme activity, genetic background, grasses, lignin, methyltransferases, missense mutation, mutants, proteins, saccharification, sequence analysis, stop codon
Modifying lignin content and composition are targets to improve bioenergy crops for cellulosic conversion to biofuels. In sorghum and other C4 grasses, the brown midrib mutants have been shown to reduce lignin content and alter its composition. Bmr12 encodes the sorghum caffeic O-methyltransferase (COMT), which catalyzes the penultimate step in monolignol biosynthesis. From an EMS-mutagenized TILLING population, four bmr12 mutants were isolated. DNA sequencing identified the 4 missense mutations in the Bmr12 coding region, which changed evolutionarily conserved amino acids Ala71Val, Pro150Leu, Gly225Asp and Gly325Ser. The previously characterized bmr12 mutants all contain premature stop codons. These mutants, along with the previously characterized bmr12-ref, represent the first allelic series of COMT mutants available in the same genetic background. The impacts of these mutants have on protein accumulation, COMT enzyme activity, Klason lignin content, lignin subunit composition and saccharification yield were determined. Gly225Asp mutant greatly reduced protein accumulation, and Pro150Leu and Gly325Ser greatly impaired COMT activity compared to wild-type (WT). All four mutants significantly reduced Klason lignin content and altered lignin composition resulting in a significantly reduced S/G ratio relative to WT, but the overall impact of these mutants were less severe than bmr12-ref. Except for Gly325Ser, which is a hypomorphic mutant, all mutants increased the saccharification yield relative to WT. These mutants represent new tools to dampen lignin content and S/G ratio, which were not possible in previous nonsense mutants, leading toward the ability to tailor lignin content and composition in the bioenergy grass sorghum.