Structures and DNA-Binding Activities of the Hinge Domains from the Structural Maintenance of Chromosomes Proteins of Pyrococcus furiosus and the Mouse Condensin Complex

Griese, Julia Johanna (2010): Structures and DNA-Binding Activities of the Hinge Domains from the Structural Maintenance of Chromosomes Proteins of Pyrococcus furiosus and the Mouse Condensin Complex. Dissertation, LMU München: Fakultät für Chemie und Pharmazie

Abstract

Structural Maintenance of Chromosomes (SMC) proteins are vital for a wide range of cellular processes including chromosome structure and dynamics, gene regulation, and DNA repair. Whereas prokaryotic genomes encode for only one SMC protein that exists as a homodimer, eukaryotes possess six different SMC proteins that form three distinct heterodimeric complexes, with the holocomplexes additionally containing several specific regulatory subunits. The prokaryotic SMC complex is required for chromosome condensation and segregation. In eukaryotes, this function is carried out by the condensin complex with SMC2 and SMC4 at its core. The complex containing SMC1 and SMC3, named cohesin, is responsible for sister chromatid cohesion during mitosis and meiosis. Cohesin is also employed in DNA double-strand break repair, whereas condensin participates in single-strand break repair. The as yet unnamed SMC5-SMC6 complex is involved in several DNA repair pathways as well as homologous recombination in meiosis. SMC proteins consist of N and C-terminal domains that fold back onto each other to create an ATPase “head” domain, connected to a central “hinge” domain via long antiparallel coiled-coils. The hinge domain mediates dimerisation of SMC proteins and binds DNA, but it is not clear to what purpose this activity serves. The aim of this work was therefore to characterise the structure and function of the SMC hinge domain in more detail. Specifically, the hinge domains of the Pyrococcus furiosus SMC protein and of mouse condensin were studied. Both their high-resolution crystal structures as well as low-resolution solution envelopes were determined, and their DNA-binding activity was analysed qualitatively and quantitatively. While the SMC hinge domain fold is largely conserved from prokaryotes to eukaryotes, functionally relevant structural differences can be observed. Most importantly, the surface charge has been almost reversed throughout evolution. The data obtained confirm that of all three eukaryotic SMC complexes, condensin is most closely related to prokaryotic SMC proteins. Both the P. furiosus and the mouse condensin hinge domain preferentially bind single-stranded DNA, but the mouse condensin hinge displays a much higher affinity than its prokaryotic counterpart, suggesting that this function has been enhanced during the course of evolution. The single-stranded DNA-binding activity might be important for the function of the condensin complex in single-strand break repair, but probably plays a different role in prokaryotes, possibly in the DNA-loading process of the prokaryotic SMC complex during replication.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Keywords: Structural Maintenance of Chromosomes (SMC), condensin, hinge domain, single-stranded DNA, X-ray crystallography
• Themengebiete: 500 Naturwissenschaften und Mathematik ; 500 Naturwissenschaften und Mathematik > 540 Chemie
• Fakultäten: Fakultät für Chemie und Pharmazie
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 25. Oktober 2010
• 1. Bericht­erstatter:in: Hopfner, Karl-Peter