ELECTRON TRANSPORT PROTEINS FROM SYNECHOCOCCUS SP. PCC 7002
Open Access
- Author:
- Nomura, Christopher
- Graduate Program:
- Biochemistry, Microbiology, and Molecular Biology
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- March 30, 2001
- Committee Members:
- Juliette T J Lecomte, Committee Member
Paul Lee Babitzke, Committee Member
Olanrewaju A Sodeinde, Committee Member
John H Golbeck, Committee Member
Donald Ashley Bryant, Committee Chair/Co-Chair - Keywords:
- heme-copper oxidase
electron transport
cyanobacteria
Synechococcus sp. PCC 7002 - Abstract:
- Cyanobacteria are photosynthetic, oxygen-evolving prokaryotes that have adapted to a wide range of ecological niches. In particular, cyanobacteria represent interesting organisms to study electron transport in because they have both photosynthetic and respiratory proteins involved in electron transport on the same membrane: the thylakoid membrane. Of particular interest to our lab is the identification and characterization of the minimal conserved number of genes responsible for coding proteins used in electron transport in cyanobacteria. In order to address this issue, a Synechococcus sp. PCC 7002 cosmid library was screened with heterologous probes made from the completely sequenced genome of the freshwater cyanobacterium, Synechocystis sp. PCC 6803. These heterologous probes were also used to screen Synechococcus sp. PCC 7002 partial genomic libraries in the cases where positive hybridizations could not be identified within the cosmid libraries. In this study, 35 open reading frames in the marine cyanobacterium Synechococcus sp. PCC 7002 have been identified and sequenced which, either, encode electron transport proteins or encode accessory proteins necessary for the assembly of these electron transport proteins based on BLAST algorithm searches. These open reading frames are represented by the following genes: ndhA, ndhI, ndhG, ndhE, ndhB, ndhC, ndhK, ndhJ, ndhD1, ndhD2, ndhD3, ndhD4, ndhF1, ndhF2, ndhF3, ndhF4, ndhF5, ndhH, ndhL, ndbA, ndbB, hypE, hoxE, hoxF, hoxU, hoxY, hypD, hoxH, hoxW, hypA, hypB, hypF, hypC, hypD, petJ1, petJ2, bcpA, ctaCI, ctaDI, ctaEI, ctaCII, ctaDII, and ctaEII. These genes putatively encode subunits for the type I NADH dehydrogenase, two type II NADH dehydrogenases, the subunits and accessory proteins for a bi-directional hydrogenase, and three putative mobile electron carriers: cytochrome c6, cytochrome c62, and BcpA, and the subunits for two different members of the heme-copper cytochrome oxidase family. Most of these genes bear the highest homology to their respective counterparts in the freshwater cyanobacterial strain Synechocystis sp. PCC 6803 and a comparison between these minimal conserved sets of genes may represent the smallest number of genes necessary for these organisms to carry out electron transport. Attempts were made to inactivate several of the electron transfer protein genes identified in this study. These include the ndhB gene, which, encodes a subunit of the type I NADH dehydrogenase, the petJ1 gene that encodes cytochrome c6, the petJ2 gene that encodes a putative second c-type cytochrome, the bcpA gene which encodes a putative blue-copper protein, the hoxH gene which encodes the large Ni containing subunit of the hydrogenase enzyme, the hoxF gene which encodes the FMN containing subunit of the hydrogenase enzyme, the ctaDI gene, which encodes the large subunit of the type I cyanobacterial cytochrome oxidase, and the ctaDII gene, which encodes the large subunit of a secondary heme-copper oxidase in Synechococcus sp. PCC 7002. This study found that the ndhB and petJ1genes are essential for the viability of Synechococcus sp. PCC 7002 and therefore could not be inactivated. The hoxH, hoxF, petJ2, bcpA, ctaDI, and ctaDII genes are all non-essential to Synechococcus sp. PCC 702 under normal growth conditions and could be inactivated. Physiological studies of the hoxH, hoxF, petJ2, and bcpA inactivated strains of Synechococcus sp. PCC 7002 revealed that there were no significant phenotypes under the conditions tested. However, evidence has been found in this study that the heme-copper oxidase enzymes encoded by the ctaCIDIEI and ctaCIIDIIEII gene clusters have significant roles in respiration, high-light tolerance and oxidative stress responses in Synechococcus sp. PCC 7002.