Unusually high mechanical stability of bacterial adhesin extender domains having calcium clamps.

To gain insight into the relationship between protein structure and mechanical stability, single molecule force spectroscopy experiments on proteins with diverse structure and topology are needed. Here, we measured the mechanical stability of extender domains of two bacterial adhesins MpAFP and MhLap, in an atomic force microscope. We find that both proteins are remarkably stable to pulling forces between their N- and C- terminal ends. At a pulling speed of 1 µm/s, the MpAFP extender domain fails at an unfolding force Fu = 348 ± 37 pN and MhLap at Fu = 306 ± 51 pN in buffer with 10 mM Ca2+. These forces place both extender domains well above the mechanical stability of many other ß-sandwich domains in mechanostable proteins. We propose that the increased stability of MpAFP and MhLap is due to a combination of both hydrogen bonding between parallel terminal strands and intra-molecular coordination of calcium ions.

Interaction of ice binding proteins with ice, water and ions.

Ice binding proteins (IBPs) are produced by various cold-adapted organisms to protect their body tissues against freeze damage. First discovered in Antarctic fish living in shallow waters, IBPs were later found in insects, microorganisms, and plants. Despite great structural diversity, all IBPs adhere to growing ice crystals, which is essential for their extensive repertoire of biological functions. Some IBPs maintain liquid inclusions within ice or inhibit recrystallization of ice, while other types suppress freezing by blocking further ice growth. In contrast, ice nucleating proteins stimulate ice nucleation just below 0 °C. Despite huge commercial interest and major scientific breakthroughs, the precise working mechanism of IBPs has not yet been unraveled. In this review, the authors outline the state-of-the-art in experimental and theoretical IBP research and discuss future scientific challenges. The interaction of IBPs with ice, water and ions is examined, focusing in particular on ice growth inhibition mechanisms.

Suspended crystalline films of protein hydrophobin I (HFBI).

Protein interfaces play an essential role in both natural and man-made materials as stabilizers, sensors, catalysts, and selective channels for ions and small molecules. Probing the molecular arrangement within such interfaces is of prime importance to understand the relation between structure and functionality. Here we report on the preparation and characterization of large area suspended crystalline films of class II hydrophobin HFBI. This small, amphiphilic globular protein readily self-assembles at the air-water interface into a 2D hexagonal lattice which can be transferred onto a holey carbon electron microscopy grid yielding large areas of hundreds of square micrometers intact hydrophobin film spun across micron-sized holes. Fourier transform analysis of low-dose electron microscopy images and selected area electron diffraction profiles reveal a unit cell dimension a=5.6±0.1nm, in agreement with reported atomic force microscopy studies on solid substrates and grazing incidence X-ray scattering experiments at the air-water interface. These findings constitute the first step towards the utilization of large-area suspended crystalline hydrophobin films as membranes for ultrapurification and chiral separation or as biological substrates for ultrafast electron diffraction.