Plant Molecular Biology

, Volume 76, Issue 3, pp 345–355

An efficient downstream box fusion allows high-level accumulation of active bacterial beta-glucosidase in tobacco chloroplasts

  • Benjamin N. Gray
  • Huijun Yang
  • Beth A. Ahner
  • Maureen R. Hanson
Article

DOI: 10.1007/s11103-011-9743-7

Cite this article as:
Gray, B.N., Yang, H., Ahner, B.A. et al. Plant Mol Biol (2011) 76: 345. doi:10.1007/s11103-011-9743-7

Abstract

Production of enzymes for lignocellulose hydrolysis in planta has been proposed as a lower-cost alternative to microbial production, with plastid transformation as a preferred method due to high foreign protein yields. An important regulator of chloroplast protein production is the downstream box (DB) region, located immediately downstream of the start codon. Protein accumulation can vary over several orders of magnitude by altering the DB region. Experiments in bacteria have suggested that these differences in protein accumulation may result from changes in translation efficiency, though the precise mechanism of DB function is not known. In this study, three DB regions were fused to the bglC ORF encoding a β-glucosidase from the thermophilic bacterium Thermobifida fusca and inserted into the tobacco (Nicotiana tabacum) plastid genome. More than a two order of magnitude of difference in BglC protein accumulation was observed, dependent on the identity of the DB fusion. Differential transcript accumulation explained some the observed differences in protein accumulation, but in addition, less 3′ degradation of bglC transcripts was observed in transgenic plants that accumulated the most BglC enzyme. Chloroplast-produced BglC was active against both pure cellobiose and against tobacco lignocellulose. These experiments demonstrate the potential utility of transplastomic plants as a vehicle for heterologous β-glucosidase production for the cellulosic ethanol industry.

Keywords

Transplastomic Beta-glucosidase Tobacco Thermobifida fusca Cellulosic ethanol Downstream box 

Supplementary material

11103_2011_9743_MOESM1_ESM.doc (67 kb)
Supplementary material 1 (DOC 67 kb)

Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Benjamin N. Gray
    • 1
    • 2
  • Huijun Yang
    • 1
  • Beth A. Ahner
    • 1
  • Maureen R. Hanson
    • 3
  1. 1.Department of Biological and Environmental EngineeringCornell UniversityIthacaUSA
  2. 2.Agrivida, Inc.MedfordUSA
  3. 3.Department of Molecular Biology and GeneticsCornell UniversityIthacaUSA

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