Protein Engineering for Photobiological Hydrogen Production

  • Development of renewable energy is essential in achieving economic growth goals and to help address the persisting world's energy crisis. Hydrogen a promising fuel source has higher energy content than oil. Here, we report for the first time (according to our knowledge) the potential application of oxygen tolerant artificial hydrogenases for the biological production of hydrogen, the up-coming alternative fuel. We created Synechocystis sp. PCC 6803 strains that photoautotrophically produce hydrogen. Transformation of wild-type Synechocystis (WT-PCC6803) was performed using an integrating or a self-replicating shuttle vector plasmid to introduce the newly engineered hydrogenase genes tolerant to oxygen, into the Synechocystis sp. PCC 6803 under the influence of the very efficient, light driven psbAII promoter or under copper repressible promoter, respectively. PCR based assays were used to screen for stable transformants. Laboratory made photobioreactor systems were established for the experimental design and data acquisition for the analysis of the Synechocystis sp. 6803 cell cultures and hydrogen production. The systems described here produced on average 15 ppm H2. We showed that H2 production was due to the enhanced expression of the artificial enzyme as untransformed microorganisms produced hydrogen closer to the background value (1ppm). These results demonstrate that it is possible to channel the energy from sunlight into hydrogen gas. Methylation pattern of the bacterial genome Abstract Methylation of DNA by the Escherichia coli DNA adenine methyltransferase (EcoDam) plays a pivot role in providing signals that influences and controls various biological processes in the bacterial cell, ranging from gene regulation and phase variation, mismatch repair, to transcription. Additionally, DNA methylation at the origin of replication is believed to play a significant role in chromosome replication and cell cycle. In the first part of this research work, we were able to analyzed and establish the relationship between the methylation of GATC sites at the origin of replication and growth phase. It was observed that the origin of replication (oriC) is kept in hemi-methylated state for nearly one-third of the cell cycle. Furthermore, we were able to determine methylation delay of DNA in the cell after replication. It was also observed that the relative activity of EcoDam reduces as cells approaches stationary phase. Additionally, we analyzed the methylation profile of the entire bacterial genome by the controlled expression of EcoDam in E. coli SCS110 cells. It was later observed that, after induction, EcoDam is capable of methylating the entire bacterial genome within 5 min. This gave a clear indication on how active EcoDam is.

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Publishing Institution:IRC-Library, Information Resource Center der Jacobs University Bremen
Granting Institution:Jacobs Univ.
Author:Martin Kangwa
Referee:Albert Jeltsch, Matthias Ullrich, Wolfgang Hilt
Advisor:Albert Jeltsch
Persistent Identifier (URN):urn:nbn:de:101:1-201305294811
Document Type:PhD Thesis
Date of Successful Oral Defense:2012/08/22
Year of Completion:2012
Date of First Publication:2012/09/24
PhD Degree:Biochemistry
School:SES School of Engineering and Science
Library of Congress Classification:T Technology / TA Engineering (General). Civil engineering (General) / TA164 Bioengineering
Call No:Thesis 2012/30

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