Interactive Crystallomic - Molecules: Nucleation, Aggregation and Crystallization - page 227

(ii) use precipitant polyethylene glycol, which has a molecular weight of
20 000 (PEG20 000) instead of commonly used salts, organic solu-
tions or low molecular weight polyethylene glycols.

Chemicals and crystallization set-up

A stock solution of BK-3 virus-like particles at a concentration of
10 mg/ml (provided by Josefina Nilsson, KI, Sweden) was used. The BK-3
particles are spherical and have a diameter of about 400 Å. A database
search on viruses (http://mmtsb.scripps.edu/viper/CRYSTALS/XTAL1/
crystals.html) confirmed that crystallization of such virus-like particles
has never previously been attempted. To check purity, SDS-PAGE was
performed. The solution was not contaminated by other membranes or
organelles since only one protein band was detected (data not shown
here). The crystallization method used was the so-called “hanging drop”
employing a reservoir of 1000 µ l in an air-tight well (Costar plate, 4 × 6

wells, Hampton Research, Laguna Niguel, CA, USA). After mixing 3 µ l

of virus-like solution (dispersed in water) with 3 µ l of reservoir, the total

volume of the drop was 6 l. The siliconized quartz cover slips — on

which the crystallization drops were created — had a diameter of 18 mm.

µ

Rationale and design of the crystallization trials

The strategy we implemented, which we call crystallomic, is an “interac-
tive protocol for crystallization.” In practical terms this means that the
composition of the reservoir (mother liquid) for a subsequent crystalliza-
tion trial ( n ), depends on what was noted immediately after the drop was
created in trial ( n 1). When the drop is created by mixing one volume of

solution of virus particles with the same volume of reservoir, there are
most often two possibilities: (a) the drop is clear and transparent, or
(b) the drop is not clear and an amorphous precipitate develops rapidly, the
drop becomes cloudy because of phase separation, or the nearly instanta-
neous formation of micro-crystals has occurred. The idea of crystallomic
is to set up several crystallization “hanging drops,” and, as long as the

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