Of spiders, bugs, and men : Structural and functional studies of proteins involved in assembly

Sammanfattning: Protein assembly enables complex machineries while being economical with genetic information. However, protein assembly also constitutes a potential threat to the host, and needs to be carefully regulated.Sulfate is a common source of sulfur for cysteine synthesis in bacteria. A putative sulfate permease CysZ from Escherichia coli appears much larger than its apparent molecular mass when analyzed by chromatography and native gel. Clearly CysZ undergoes homo-oligomerization. Using isothermal titration calorimetry, we confirmed that CysZ binds to its putative substrate sulfate, and also sulfite with higher affinity. CysZ-mediated sulfate transport—in both E. coli whole cells and proteoliposomes—was inhibited in the presence of sulfite, indicating a feedback inhibition mechanism.Proteus mirabilis is a Gram-negative bacterium causing urinary tract infections. Its simultaneous expression of multiple fimbriae enables colonization and biofilm formation. Fimbriae are surface appendages assembled from protein subunits, with distal adhesins specifically recognizing host-cell receptors. We present the first three structures of P. mirabilis fimbrial adhesins. While UcaD and AtfE adopt the canonical immunoglobulin-like fold, MrpH has a previously unknown fold. The coordination of Zn or Cu ion by three conserved histidine residues in MrpH is required for MrpH-dependent biofilm formation.Spider silk is an assembly of large proteins called spidroins. The N-terminal domain (NT) of spidroins senses the pH decrease along the silk spinning gland, and transits from monomer to dimer. A locked NT dimer interlinks spidroin molecules into polymers. We identified a new asymmetric dimer form of NT by x-ray crystallography. With additional evidence from small angle x-ray scattering (SAXS), we propose the asymmetric dimer as a common intermediate of NT in silk formation.Alzheimer’s disease is a life-threatening dementia, where aggregation-prone Aβ peptides self-assemble into amyloid fibrils. Bri2 BRICHOS is a molecular chaperone that efficiently delays Aβ fibrillation, and protects the region of its pro-protein with high β-propensity from aggregation. Combining SAXS and microscale thermophoresis data, we confirmed binding between Bri2 BRICHOS and its native client peptide. Using site-directed mutagenesis, we showed that three conserved tyrosine residues in Bri2 BRICHOS are important for its anti-Aβ fibrillation activity.