PRI HOTSCORE. Web server for the prediction of amino acid
hotspots in RNA–protein interfaces using in silico alanine
scanning mutagenesis (https://pri-hotscore.labs.vu.nl).
RCRANE. A plugin that facilitates semiautomated building of
RNA structures within COOT (please refer to https://
pylelab.org/software; RCrane is included with COOT
0.7 or newer).
RIBER/DIBER. A software suite for crystal content analysis in
the studies of protein–nucleic acid complexes (http://
diber.iimcb.gov.pl/).
RNA 3D MOTIF ATLAS. A comprehensive and representative
collection of internal and hairpin loop RNA threedimensional motifs extracted from representative sets of
RNA three-dimensional structures (http://rna.bgsu.edu/
rna3dhub/motifs).
RNA BRICKS. A database of RNA three-dimensional structure motifs and their contacts, both with themselves and
with proteins. The database provides structure-quality
score annotations and tools for three-dimensional RNA
structure searches and comparison (http://iimcb.
genesilico.pl/rnabricks/).
RNA FRABASE 2.0. An engine with database to search the
three-dimensional fragments within three-dimensional
RNA structures using the sequence(s) and/or secondary
structure(s) as input (http://rnafrabase.cs.put.poznan.pl/).
SWISS-MODEL. A fully automated protein structure homology modeling server (https://swissmodel.expasy.org/).
5.
32 P-labeling represents a commonly used method for making
RNAs detectable in the electrophoretic mobility shift assay in
labs where radioisotope use in permitted. Labeled RNA probes
can be prepared by in vitro transcription using [α32 P]NTPs,
whereas oligoribonucleotide or 100–200-nucleotide purified
RNA can be end-labeled using [
32 P]cytidine 3
0 -5
0 -bis(phosphate) or [γ32 P]ATP. Alternatively, RNAs can be covalently
labeled with a fluorophore in labs where radioisotope use in not
permitted. Unlabeled RNAs can be used and detected by postelectrophoretic staining with chromophores or fluorophores
that bind to the RNA.
6. Sample concentration is a significant crystallization variable.
Samples that are too concentrated can result in amorphous
precipitate whereas samples that are too dilute can result in
clear drops. The PCT™ (pre-crystallization test; Hampton
Research) can be used to determine the appropriate protein
concentration for crystallization screening to reduce the number of precipitate and clear results and increase the likelihood of
444
Andrew P. Turnbull and Xiaoqiu Wu
hotspots in RNA–protein interfaces using in silico alanine
scanning mutagenesis (https://pri-hotscore.labs.vu.nl).
RCRANE. A plugin that facilitates semiautomated building of
RNA structures within COOT (please refer to https://
pylelab.org/software; RCrane is included with COOT
0.7 or newer).
RIBER/DIBER. A software suite for crystal content analysis in
the studies of protein–nucleic acid complexes (http://
diber.iimcb.gov.pl/).
RNA 3D MOTIF ATLAS. A comprehensive and representative
collection of internal and hairpin loop RNA threedimensional motifs extracted from representative sets of
RNA three-dimensional structures (http://rna.bgsu.edu/
rna3dhub/motifs).
RNA BRICKS. A database of RNA three-dimensional structure motifs and their contacts, both with themselves and
with proteins. The database provides structure-quality
score annotations and tools for three-dimensional RNA
structure searches and comparison (http://iimcb.
genesilico.pl/rnabricks/).
RNA FRABASE 2.0. An engine with database to search the
three-dimensional fragments within three-dimensional
RNA structures using the sequence(s) and/or secondary
structure(s) as input (http://rnafrabase.cs.put.poznan.pl/).
SWISS-MODEL. A fully automated protein structure homology modeling server (https://swissmodel.expasy.org/).
5.
32 P-labeling represents a commonly used method for making
RNAs detectable in the electrophoretic mobility shift assay in
labs where radioisotope use in permitted. Labeled RNA probes
can be prepared by in vitro transcription using [α32 P]NTPs,
whereas oligoribonucleotide or 100–200-nucleotide purified
RNA can be end-labeled using [
32 P]cytidine 3
0 -5
0 -bis(phosphate) or [γ32 P]ATP. Alternatively, RNAs can be covalently
labeled with a fluorophore in labs where radioisotope use in not
permitted. Unlabeled RNAs can be used and detected by postelectrophoretic staining with chromophores or fluorophores
that bind to the RNA.
6. Sample concentration is a significant crystallization variable.
Samples that are too concentrated can result in amorphous
precipitate whereas samples that are too dilute can result in
clear drops. The PCT™ (pre-crystallization test; Hampton
Research) can be used to determine the appropriate protein
concentration for crystallization screening to reduce the number of precipitate and clear results and increase the likelihood of
444
Andrew P. Turnbull and Xiaoqiu Wu