Details

Autor(en) / Beteiligte

• Hart, Benjamin R

Titel

Expanded functionality of the bacterial global regulator Lrp

Ort / Verlag

ProQuest Dissertations & Theses

Erscheinungsjahr

2010

Quelle

ProQuest Dissertations & Theses A&I

Beschreibungen/Notizen

• Predicting gene regulation from genome sequences is an important technique for understanding bacteria that cannot currently be grown in the laboratory. This approach involves extrapolation from a well-characterized bacterium. Several assumptions are made when using this technique; key among these is that sequence-conserved transcription factors, target genes, and binding sites for these transcription factors upstream of the target genes together imply conservation of the regulation. However the level of conservation necessary for accurate predictions has not been defined. Previous studies have illustrated that the Leucine Responsive Regulatory Protein (Lrp) orthologs from Escherichia coli and Proteus mirabilis have only partially-conserved regulatory effects despite 98% overall amino acid sequence identity and complete conservation of the DNA binding helix-turn-helix domain. Studies described here reveal that these regulatory differences are associated with previously-unappreciated but fundamental functional differences between the Lrp orthologs. These studies are particularly important for predicting regulation from genome sequence as Lrp is a global regulator that, in E. coli, directly controls over 200 genes. The first manuscript, prepared for the Journal of Bacteriology, focuses on the amino acid coregulators of Lrp that, for E. coli, were only previously known to include leucine and alanine. This study revealed that methionine, isoleucine, histidine, and threonine also have significant coregulatory effects. In addition, modest differences between Lrp orthologs were observed in response to some amino acids. The second manuscript focuses on the role of the N-terminal tail of the Lrp protein. This unstructured tail is a region containing many of the sequence differences between Lrp orthologs. Through the generation of hybrid proteins, this study demonstrates that the N-terminal tails contribute to differences in transcription, and DNA binding between E. coli and P. mirabilis Lrp. Together, these results suggest that even overall sequence identity of 98% is insufficient to allow regulatory extrapolation in the absence of fairly detailed understanding of the regulatory protein.