Photon Factory Activity Report 2019 #37 (2020)

BL-6A/2018G526

Attempt to observe the structure of schizophyllan monomer by Small Angle X-ray
Scattering and Molecular Dynamics simulation
Yoshitaka Matsumura* and Masaki Kojima
School of Life Sciences, Tokyo University of Pharmacy and Life Sciences, 1432-1 Horinouchi,Hachioji, Tokyo 192-0392, Japan

2 Experiments
All the SAXS experiments were carried out at thebeamline 6A using a PILATUS detector. The
measurements were repeated under equilibrium at 25℃
with the sample-to-detector distance of ca. 0.5 m. For theGuinier analyses, both single and multiple exponential
approximations were adopted [4, 5]. When irregularaggregates were contaminated in the sample solution, themultiple exponential approximation is very convenientand useful to obtain the structural properties of the major
component in the mixture as was previously reported forother samples [5]. Bovine serum albumin and hen egg
white lysozyme were employed as a reference for thecalibration. Before the SAXS measurement, the solutionof schizophyllan was sonicated for at least 200 hours. The
sample concentration was less than 10 mg/mL. Molecular
dynamics simulation was performed by Amber program
package [6].
3 Results and Discussion
Figure 1 shows the Guinier plot (a) and the Kratky plot(b) of schizophyllan upon sonication. The Guinier plot
indicated that the schizophyllan molecule was in a multidispersed state, and the obvious peak in the Kratky plotsuggested a globular shape. The estimated radius ofgyration (Rg) of the major component was almost thesame (ca. 10 Å) in the Guinier and Kratky plots at several
concentrations. Compared of its molecular weight fromthe Guinier plots with those reported previously, themajor component in the current study could exist as amonomer [3]. The plausible structure was predicted by
MD simulation using Amber with the molecular weight
and the primary structure of the monomeric form
obtained from the Guinier analyses. The resultantstructure by MD simulation is shown in Figure 2 (a) and(b), which respectively show the stick model and the traceof the backbone. The backbone trace depicted a helicalstructure. Figure 2 (c) shows a coarse-grained structure of
the major component of the schizophyllan constructed byab initio modeling [7]. When the MD structure wasaligned along the same inertial axis of the ab initio model,
both structures were well superimposed (Figure 2 (d)).
This indicates that the MD structure satisfied the SAXS
structural information.
1-1-3-5-7-9-11-13-15

0.000250.0002I*h*hln(I)

1 Introduction
Schizophyllan, a β-glucan derived from Schizophyllum
commune, has been approved as an anticancer agent
because it exhibits antitumor activity. The X-ray crystal
structure analysis revealed that it takes a triple helixstructure in the native state [1, 2] . On the other hand, it
has been reported that the sonication of schizophyllan
reduced the molecular weight and caused a structuralchange from trimer to monomer, resulting in loss ofanticancer activity [3]. So, we aimed to observe andvisualize the three-dimensional structure of themonomeric schizophyllan produced upon sonication bysmall angle X-ray scattering (SAXS) and molecular
dynamics (MD) simulation.

0.000150.0001◯ Experimental data
− Guinier fitting

0.00005
00 0.01 0.02 0.03 0.04 0.05 0.06 0.07
h2 (Å-2)

00.1

0.2 0.3h (Å-1)

0.4

0.5

Figure 1. Guinier plot (a:left) and Kratky plot (b: right)of schizophyllan
(a)

(b)

(c)

(d)

Figure 2. (a): Stick representation of the predictedstructure from MD simulation, (b): The backbone traceof the structure (a), (c): coarse-grained structure from ab
initio modeling, (d): superimposed structures (a) and (c).
References[1] Y. Takahashi, et al. Rep. Prog. Polym. Phys. Jpn. 27,767, (1984)[2] D. B. Kony, et al. Biophys. J. 93, 442-455, (2007)[3] T. Yanaki, et al. Biophys. Chem. 17, 337-342, (1983)[4] A. Guinier, and G. Fournet, Wiley New York, (1955)[5] Y. Matsumura, et al. Biophys. Chem. 175-176, 39-46,(2013)[6] R. Salomon-Ferrer, et al. WIREs Comput. Mol. Sci. 3,198-210, (2013)[7] D. Franke, and D. I. Svergun, J. Appl. Cryst. 42, 342346, (2009)
OutcomeSpeaker, The 6th International Symposium on
Bioimaging, The 28th Annual Meeting of Bioimaging
Society, 2019* matsumuy@toyaku.ac.jp