5XX7

Hetero-micro-seeding: Crystal structure of BPTI-[5,55]C14GA38I variant using micro-seeds from -C14GA38I variant

Summary for 5XX7

• Entry DOI: 10.2210/pdb5xx7/pdb
• Descriptor: Pancreatic trypsin inhibitor, SULFATE ION (3 entities in total)
• Functional Keywords: hydrolase inhibitor, hydrolase
• Biological source: Bos taurus (Bovine)
• Total number of polymer chains: 2
• Total formula weight: 13106.84

Authors

Islam, M.M. (deposition date: 2017-07-01, release date: 2018-07-04, Last modification date: 2024-11-13)

Primary citation

Islam, M.M.,Kuroda, Y.
A hetero-micro-seeding strategy for readily crystallizing closely related protein variants
Biochem. Biophys. Res. Commun., 493:504-508, 2017

PubMed Abstract: Protein crystallization remains difficult to rationalize and screening for optimal crystallization conditions is a tedious and time consuming procedure. Here, we report a hetero-micro-seeding strategy for producing high resolution crystals of closely related protein variants, where micro crystals from a readily crystallized variant are used as seeds to develop crystals of other variants less amenable to crystallization. We applied this strategy to Bovine Pancreatic Trypsin Inhibitor (BPTI) variants, which would not crystallize using standard crystallization practice. Out of six variants in our analysis, only one called BPTI-[5,55]A14G formed well behaving crystals; and the remaining five (A14GA38G, A14GA38V, A14GA38L, A14GA38I, and A14GA38K) could be crystallized only using micro-seeds from the BPTI-[5,55]A14G crystal. All hetero-seeded crystals diffracted at high resolution with minimum mosaicity, retaining the same space group and cell dimension. Moreover, hetero-micro-seeding did not introduce any biases into the mutant's structure toward the seed structure, as demonstrated by A14GA38I structures solved using micro-seeds from A14GA38G, A14GA38L and A14GA38I. Though hetero-micro-seeding is a simple and almost naïve strategy, this is the first direct demonstration of its workability. We believe that hetero-micro-seeding, which is contrasting with the popular idea that crystallization requires highly purified proteins, could contribute a new tool for rapidly solving protein structures in mutational analysis studies.

PubMed: 28870811
DOI: 10.1016/j.bbrc.2017.08.161

Experimental method

X-RAY DIFFRACTION (1.38 Å)