Multiple binding modes of a moderate ice-binding protein from a polar microalga

Abstract

Ice-binding proteins (IBPs) produced by cold-tolerant organisms interact with ice and strongly control crystal growth. The molecular basis for the different magnitudes of activity displayed by various IBPs (moderate and hyperactive) has not yet been clarified. Previous studies questioned whether the moderate activity of some IBPs relies on their weaker binding modus to the ice surface, compared to hyperactive IBPs, rather than relying on binding only to selected faces of the ice crystal. We present the structure of one moderate IBP from the sea-ice diatom Fragilariopsis cylindrus (fcIBP) as determined by X-ray crystallography and investigate the protein's binding modes to the growing ice-water interface using molecular dynamics simulations. The structure of fcIBP is the IBP-1 fold, defined by a discontinuous β-solenoid delimitated by three faces (A, B and C-faces) and braced by an α-helix. The fcIBP structure shows capping loops on both N- and C-terminal parts of the solenoid. We show that the protein adsorbs on both the prism and the basal faces of ice crystals, confirming experimental results. The fcIBP binds irreversibly to the prism face using the loop between the B and the C-faces, involving also the B-face in water immobilization despite its irregular structure. The α-helix attaches the protein to the basal face with a partly reversible modus. Our results suggest that fcIBP has a looser attachment to ice and that this weaker binding modus is the basis to explain the moderate activity of fcIBP.

Ice‐binding proteins and the ‘domain of unknown function’ 3494 family

“…This binding pattern to ice crystal planes was confirmed also for the moderate IBP isoform 11 from the sea‐ice diatom Fragilariopsis cylindrus ( fc IBP11) by laser confocal fluorescence microscopy of a single ice crystal in a solution of the fluorescently labelled protein . In addition to these experimental methods, MD simulations of fc IBP11 indicated that the protein can, at least partly, bind the primary prism and basal faces, despite its moderate activity .…”

“…A) being the most representative (~ 68%). Almost all characterized DUF3494 IBPs exhibit this simple architecture . Generally, single‐DUF3494 IBPs also contain an N‐terminal signal peptide, suggesting that these proteins are secreted into the environment near the cells or accumulate in the membrane .…”

“…Currently, nine crystal structures have been reported for DUF3494 IBP family members: Tis IBP6 , Tis IBP8 , Leucosporidium sp. AY30 IBP ( Le IBP) , Ff IBP , Col IBP , IBPv , Sf IBP_1 , Efc IBP and fc IBP11 . All these proteins present the IBP‐1 fold that consists of a discontinuous right‐handed β‐solenoid with a triangular cross‐section formed by three parallel β‐sheets (faces a, b and c), and an α helix that runs along the a‐face, parallel to the main axis of the protein (Fig.…”

“…Originally, this group was found only in eukaryotic IBPs (e.g. Le IBP, Tis IBP, fc IBP11), but the bacterial Sf IBP_1's capping structure was elucidated and shown to contain a superficially similar structure, albeit with very different residue contacts holding the loops together . The final group currently includes only Efc IBP, which does not contain any capping structure (Fig.…”

“…Far more consistent between DUF3494 IBPs, it usually folds as a small loop that cuts across the helix end and blocks the hydrophobic core. An exception is represented by fc IBP11, whose C‐terminal capping sports a distinctly larger loop .…”