Tartrate Chirality Determines Thaumatin Crystal Habit

Abstract

A major challenge in structural biology is to produce high-quality protein crystals for X-ray diffraction. Currently, proteins are crystallized by trial and error, often in multicomponent solutions with chiral precipitants. As proteins are chiral molecules, we hypothesized that the chirality of the precipitants may affect crystallogenesis. To test this hypothesis, we crystallized thaumatin, an intensely sweet globular protein, with the three stereoisomers (l-, d-, and meso-) of tartaric acid. We find three different crystal habits and crystal packings; the three stereoisomers interact with the protein at different sites. All three precipitants produce high-quality crystals from which atomic resolution (∼1 Å) structures were obtained. Our findings suggest that stereospecific interactions with precipitants are important in protein crystal formation and should be controlled when crystallizing proteins for structure determination.

Protein crystallization in short-peptide supramolecular hydrogels: a versatile strategy towards biotechnological composite materials

“…This can be explained by (i) the physical properties of the hydrogel which eliminates sedimentation, convection current 7 and acts as impurity filter media, 8 and (ii) their molecular influence, as hydrogel fibers can interact directly with protein molecules, 9,10 having a final composite formed by polymeric fibers of agarose, 11 silica, 12,13 and PEG-based hydrogels, 14 incorporated within the crystal lattices of protein crystals. 26 Similarly, we have also found a chiral influence on the crystal growth and quality of model proteins using homochiral peptidic hydrogels. 18 Considering this, very recently we have shown that novel short-peptide supramolecular hydrogels serve as convective free media to grow protein crystals of high quality.…”

Atomic structure of recombinant thaumatin II reveals flexible conformations in two residues critical for sweetness and three consecutive glycine residues

“…These residues were located posterior to the cleft-containing region. Four charged residues (Glu35, Asp55, Arg171 and Arg200) were also identified in plant thaumatin I (PDB code 2vhk [6], PDB code 3ald, [7]) and recombinant thaumatin I (PDB code 3al7, [7]). Notably, we could successfully identify two conformations for G143 (occupancy was 0.58 for G143 (A) and 0.42 for G143 (B), respectively) (Fig.…”

“…Since structural information on the charge distribution and protonation state of hydrogen atoms would be useful for understanding the elicitation of the sweetness of thaumatin as well as the interaction with sweet receptors, high-resolution x-ray crystal analysis and neutron crystallographic studies have been conducted [6e10]. Atomic resolution structures with hydrogen atoms of recombinant thaumatin I (1.0e1.1 Å) as well as purified plant thaumatin I (0.94 Å) demonstrated that electron density maps of some residues were significantly improved to clarify subtle structural variations among thaumatin variants [6,7]. Furthermore, comparisons of structures at atomic resolution revealed the large disulfide-rich region in domain II to be sensitive to a pH change [8].…”

Custom-tailored adsorbers: A molecular dynamics study on optimal design of ion exchange chromatography material

“…9 Various studies on the influence of ligand density on binding behavior have been 10 conducted in the past. Wu and Walters [2] found in a study on ion exchange 11 isocratic elution experiments with two proteins that the Z number, which de- 12 scribes the number of interaction binding sites of the protein with the ligand 13 surface, increased as the ligand density increased. This behavior is comprehen-14 sible from a molecular point of view, since more binding sites of the protein are 15 within the reach of (electrostatic) interaction with the ligands.…”