High-throughput Automated System for Crystallizing Membrane Proteins in Lipidic Mesophases.

Abstract

A high-throughput robotic system has been developed to enable the automatic handling of nanoliter volumes of highly viscous biomaterials for crystallizing membrane proteins using lipidic mesophases. The in meso method introduced a few years ago has produced crystal structures of a number of important membrane proteins. The bottleneck to achieve high throughput automation of this, so-called in meso method, is the handling of nanoliter volumes of the highly viscous cubic phase as part of the crystallization process. The cubic phase sticks to everything that it has contact with and has a tendency to disable dispensing robotic tools. In this paper, we discuss the factors that influence the successful and automatic delivery of nanoliter volumes of the cubic phase. A creative cubic-phase-based coordinate measuring mechanism is presented for controlling the dispensing distance of the cubic phase which is critically important for the successful performance of the system. A mathematical model describing the cubic-phase delivery is proposed and verified.We also present the optimization of liquid handling parameters for the successful and automatic delivery of different precipitant solutions. The performance characteristics of the robotic system in terms of accuracy and reproducibility of delivering nano volumes of highly viscous biomaterials and micro volumes of different precipitant solutions are reported.