Biochemical characterization of essential cell division proteins FtsX and FtsE that mediate peptidoglycan hydrolysis by PcsB in Streptococcus pneumoniae
| Autor: | Kevin E. Bruce, Britta E. Rued, Amy L. Davidson, Malcolm E. Winkler, Cynthia V. Stauffacher, Ruchika Bajaj |
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| Jazyk: | angličtina |
| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
bacterial cell division Cell division ATPase 030106 microbiology Detergents ATP-binding cassette transporter Cell Cycle Proteins Peptidoglycan Biology medicine.disease_cause Microbiology ABC transporter‐like proteins FtsEX:PcsB complex 03 medical and health sciences chemistry.chemical_compound peptidoglycan (PG) hydrolases Bacterial Proteins medicine Extracellular Escherichia coli Integral membrane protein Micelles Original Research Membrane Proteins Protein Structure Tertiary 030104 developmental biology Streptococcus pneumoniae chemistry Biochemistry Cytoplasm biology.protein ATP-Binding Cassette Transporters pneumococcus Cell Division Protein Binding |
| Zdroj: | MicrobiologyOpen |
| ISSN: | 2045-8827 |
| Popis: | The FtsEX:PcsB complex forms a molecular machine that carries out peptidoglycan (PG) hydrolysis during normal cell division of the major respiratory pathogenic bacterium, Streptococcus pneumoniae (pneumococcus). FtsX is an integral membrane protein and FtsE is a cytoplasmic ATPase that together structurally resemble ABC transporters. Instead of transport, FtsEX transduces signals from the cell division apparatus to stimulate PG hydrolysis by PcsB, which interacts with extracellular domains of FtsX. Structural studies of PcsB and one extracellular domain of FtsX have recently appeared, but little is known about the biochemical properties of the FtsE ATPase or the intact FtsX transducer protein. We report here purifications and characterizations of tagged FtsX and FtsE proteins. Pneumococcal FtsX‐GFP‐His and FtsX‐His could be overexpressed in Escherichia coli without toxicity, and FtsE‐His remained soluble during purification. FtsX‐His dimerizes in detergent micelles and when reconstituted in phospholipid nanodiscs. FtsE‐His binds an ATP analog with an affinity comparable to that of ATPase subunits of ABC transporters, and FtsE‐His preparations have a low, detectable ATPase activity. However, attempts to detect complexes of purified FtsX‐His, FtsE‐His, and PcsB‐His or coexpressed tagged FtsX and FtsE were not successful with the constructs and conditions tested so far. In working with nanodiscs, we found that PcsB‐His has an affinity for charged phospholipids, mediated partly by interactions with its coiled‐coil domain. Together, these findings represent first steps toward reconstituting the FtsEX:PcsB complex biochemically and provide information that may be relevant to the assembly of the complex on the surface of pneumococcal cells. |
| Databáze: | OpenAIRE |
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