Domain-swapped structure of the potent antiviral protein griffithsin and its mode of carbohydrate binding

Abstract

The crystal structure of griffithsin, an antiviral lectin from the red alga Griffithsia sp., was solved and refined at 1.3 A resolution for the free protein and 0.94 A for a complex with mannose. Griffithsin molecules form a domain-swapped dimer, in which two beta strands of one molecule complete a beta prism consisting of three four-stranded sheets, with an approximate 3-fold axis, of another molecule. The structure of each monomer bears close resemblance to jacalin-related lectins, but its dimeric structure is unique. The structures of complexes of griffithsin with mannose and N-acetylglucosamine defined the locations of three almost identical carbohydrate binding sites on each monomer. We have also shown that griffithsin is a potent inhibitor of the coronavirus responsible for severe acute respiratory syndrome (SARS). Antiviral potency of griffithsin is likely due to the presence of multiple, similar sugar binding sites that provide redundant attachment points for complex carbohydrate molecules present on viral envelopes.

Figure 1

Comparison of the Sequences of Griffithsin and Related Lectins (A) Structure-based sequence comparison of griffithsin and eight other lectins with a similar fold. Residues that are strictly conserved among all nine proteins are shown on red background, and those of a similar character are highlighted in yellow. Residues that are strictly conserved among the other lectins but not in griffithsin are highlighted in green in the former and magenta in the latter. (B) Structure-based sequence alignment of the three β sheets (blades) in griffithsin. Conserved residues that interact directly with the bound carbohydrates are highlighted in red, whereas conserved residues that do not make such contacts are shown on magenta background.

Figure 2

A Ribbon Cα Trace of a Griffithsin Dimer, Based on the Coordinates of the His-Tagged Version of the Protein Molecule A is colored blue, and molecule B green, with the N-terminal extension that resulted from the cloning procedure colored red.

Figure 3

Superposition of the Backbone Tracing of Griffithsin and Other Related Lectins A compact domain of griffithsin was defined as residues A1–A18 and B19–B121. The following colors were used: griffithsin, red; artocarpin (1vbo), blue; heltuba (1c3m), magenta; parkia lectin (1zgs), yellow; calsepa (1ouw), pink; banana lectin (2bmz), black; jacalin (1ugw), violet; AHL (1toq), brown; MPA (1jot), green.

Figure 4

Mannose Binding Sites 1–3, Created Principally by Molecule A of Griffithsin The omitmap F_o − F_c electron density map was calculated at 1.8 Å resolution, based on the final coordinates of the structure of crystal 1 refined after the removal of the mannoses, and it was contoured at the 2.7 σ level.

Figure 5

Details of the Interactions between Man3 and Griffithsin The locations and lengths of hydrogen bonds are representative for all six principal carbohydrate binding sites.