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Fibrin forms the structural scaffold of blood clots and has great potential for biomaterial applications. Creating recombinant expression systems of fibrinogen, fibrin’s soluble precursor, would advance the ability to construct mutational libraries that would enable structure–function studies of fibrinogen and expand the utility of fibrin as a biomaterial. Despite these needs, recombinant fibrinogen expression systems, thus far, have relied on the time-consuming creation of stable cell lines. Here we present tests of a transient fibrinogen expression system that can rapidly generate yields of 8–12 mg/L using suspension HEK Expi293TM cells. We report results from two different plasmid systems encoding the fibrinogen cDNAs and two different transfection reagents. In addition, we describe a novel, affinity-based approach to purifying fibrinogen from complex media such as human plasma. We show that using a high-affinity peptide which mimics fibrin’s knob ‘A’ sequence enables the purification of 50–75% of fibrinogen present in plasma. Having robust expression and purification systems of fibrinogen will enable future studies of basic fibrin(ogen) biology, while paving the way for the ubiquitous use of fibrin as a biomaterial. 

Fluorinated 5-hydroxytryptophans (F n -5HOWs) were synthesized in gram scale quantities and incorporated into a β-hairpin peptide and the protein azurin. The redox-active F n -5HOWs exhibit unique radical spectroscopic signatures that expand the function of 5HOW as probes for biological electron transfer. 

Previous comparative kinetic isotope effects have inferred an allosteric site for fatty acids and their derivatives that modulates substrate selectivity in 15‐lipoxygenases. Hydrogen–deuterium exchange also previously revealed regionally defined enhanced protein flexibility, centred at helix α2 – a gate to the substrate entrance. Direct evidence for allosteric binding and a complete understanding of its mechanism remains elusive. In this study, we examine the binding thermodynamics of the fatty acid mimic, oleyl sulfate (OS), with the monomeric model plant 15‐LOX, soybean lipoxygenase (SLO), using isothermal titration calorimetry. Dynamic light scattering and differential scanning calorimetry rule out OS‐induced oligomerization or structural changes. These data provide evidence that the fatty acid allosteric regulation of SLO is controlled by the dynamics of helix α2.