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Title: Protein degradation in Mycobacterium smegmatis
Authors: Knipfer, Nancianne
Keywords: Biochemistry.
Molecular biology.
Issue Date: 1998
Publisher: ProQuest Dissertations & Theses
Citation: Source: Dissertation Abstracts International, Volume: 59-06, Section: B, page: 2735.;Advisors: Thomas E. Shrader.
Abstract: Within the cytoplasm, protein degradation proceeds predominately via three classes of ATP-dependent proteases: Lon, Clp and the proteasome. A survey of the completed bacterial genomes revealed that bacteria contain multiple ATP-dependent proteases but that the distribution of these proteases is highly varied between bacterial species. In contrast, eukaryotic cells depend almost exclusively on the 26S proteasome for cytoplasmic protein degradation. Recently, a homologue of the 20S proteasome was identified in Mycobacterium leprae suggesting mycobacteria like eukaryotes, may depend exclusively on the proteasome for cytoplasmic protein degradation. The non-pathogenic, fast growing Mycobacterium smegmatis was selected as a model mycobacterial cell to study protein degradation. A novel method of positive and negative selection was developed to inactivate the genes encoding the M. smegmatis 20S proteasome. The deletion experiments revealed that in contrast to eukaryotic cells, M. smegmatis cells lacking 20S proteasome genes were viable and were phenotypically indistinguishable from wild type cells and suggested that M. smegmatis cells contain additional ATP-dependent proteases. Indeed a homolog of the Lon protease was identified in M. smegmatis. To assess the functional importance of the Lon protease, the lon coding sequences were removed from the chromosome of both a wild type and proteasome disrupted strain. In contrast to either E. coli lon mutants or hslUV mutants, M. smegmatis lon prcBA double mutants were not impaired in the degradation of abnormal proteins implying additional proteases with broad substrate specificity exist within the cell. Subsequently, our work and the M. tuberculosis genome project revealed that M. smegmatis and M. tuberculosis each contain one or more ClpP homologs. Based on the demonstration that M. tuberculosis contains two clpP homologs, we propose that unlike the E. coli Clp protease, the mycobacterial Clp protease may display broad substrate specificity. Overall, we revealed that M. smegmatis contains homologs of the proteasome, Lon and the Clp protease and demonstrated that the functions of these proteases are not conserved between M. smegmatis and E. coli.
Appears in Collections:Albert Einstein College of Medicine: Doctoral Dissertations

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