Biological activity differences between TGF-β1 and TGF-β3 correlate with differences in the rigidity and arrangement of their component monomers

[Image: see text] TGF-β1, -β2, and -β3 are small, secreted signaling proteins. They share 71–80% sequence identity and signal through the same receptors, yet the isoform-specific null mice have distinctive phenotypes and are inviable. The replacement of the coding sequence of TGF-β1 with TGF-β3 and TGF-β3 with TGF-β1 led to only partial rescue of the mutant phenotypes, suggesting that intrinsic differences between them contribute to the requirement of each in vivo. Here, we investigated whether the previously reported differences in the flexibility of the interfacial helix and arrangement of monomers was responsible for the differences in activity by generating two chimeric proteins in which residues 54–75 in the homodimer interface were swapped. Structural analysis of these using NMR and functional analysis using a dermal fibroblast migration assay showed that swapping the interfacial region swapped both the conformational preferences and activity. Conformational and activity differences were also observed between TGF-β3 and a variant with four helix-stabilizing residues from TGF-β1, suggesting that the observed changes were due to increased helical stability and the altered conformation, as proposed. Surface plasmon resonance analysis showed that TGF-β1, TGF-β3, and variants bound the type II signaling receptor, TβRII, nearly identically, but had small differences in the dissociation rate constant for recruitment of the type I signaling receptor, TβRI. However, the latter did not correlate with conformational preference or activity. Hence, the difference in activity arises from differences in their conformations, not their manner of receptor binding, suggesting that a matrix protein that differentially binds them might determine their distinct activities

Crystal structure of oxygen-evolving photosystem II at a resolution of 1.9 Å

Photosystem II is the site of photosynthetic water oxidation and contains 20 subunits with a total molecular mass of 350 kDa. The structure of photosystem II has been reported at resolutions from 3.8 to 2.9 angstrom. These resolutions have provided much information on the arrangement of protein subunits and cofactors but are insufficient to reveal the detailed structure of the catalytic centre of water splitting. Here we report the crystal structure of photosystem II at a resolution of 1.9 angstrom. From our electron density map, we located all of the metal atoms of the Mn(4)CaO(5) cluster, together with all of their ligands. We found that five oxygen atoms served as oxo bridges linking the five metal atoms, and that four water molecules were bound to the Mn(4)CaO(5) cluster; some of them may therefore serve as substrates for dioxygen formation. We identified more than 1,300 water molecules in each photosystem II monomer. Some of them formed extensive hydrogen-bonding networks that may serve as channels for protons, water or oxygen molecules. The determination of the high-resolution structure of photosystem II will allow us to analyse and understand its functions in great detail