Association kinetics of wild- and mutant-type Ynd1p in relation to quality of grown crystals.
The intermolecular interaction and association dynamics of the Ynd1p protein were investigated using dynamic and time-resolved static light scattering measurements. The mutual diffusion coefficients of wild- and mutant-type (a single amino acid substitution) Ynd1p monomer were measured in 50 mM 2-(N-morpholino)ethanesulfonic acid (MES) buffer with 5 mM MnCl2 and 7.5% (v/v) ethylene glycol. Both translational diffusion coefficients at a zero protein concentration were (40.3 +/- 0.2) x 10(-12) m2/s at 20 degrees C and a pH of 7.0, so the hydrodynamic radius of the monomers was 4.1 +/- 0.1 nm. The measured intermolecular interaction between monomers, however, showed that the mutant-type Ynd1p had a stronger attractive force. Time-resolved static light scattering measurements showed that the association of mutant-type Ynd1p yielded a larger number of aggregates than that of wild-type Ynd1p. The time dependence of aggregate gyration radius differed between the two types. Fractal dimension analysis using scattering intensity data suggested that the inner structure of the aggregates changed from loose to rigid with time. Although this phenomenon is common for wild and mutant types, the differences in the number of aggregates yielded in the initial stages and in the intermolecular interaction affected the quality of the final grown crystals. That is, single crystals of Ynd1p grew in the mutant-type protein solution and polycrystals of Ynd1p grew in the wild-type protein solution.
Onuma
Kazuo
K
Institute for Human Science and Biomedical Engineering, National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Central 6, Tsukuba, Ibaraki 305-8566, Japan. k.onuma@aist.go.jp
Watanabe
Akiko
A
Kanzaki
Noriko
N
Kubota
Tomomi
T
D001081:Apyrase / Q000737:chemistry* / Q000235:genetics*; D003460:Crystallization; D007700:Kinetics; D008027:Light; D008970:Molecular Weight; D009154:Mutation; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics*; D012542:Scattering, Radiation; D014961:X-Ray Diffraction
D029701:Saccharomyces cerevisiae Proteins; D001081:Apyrase; C506070:Ynd1 protein, S cerevisiae