Experimental models of protein-RNA interaction: isolation and analyses of tRNA(Phe) and U1 snRNA-binding peptides from bacteriophage display libraries.
Peptides that bind either U1 small nuclear RNA (U1 snRNA) or the anticodon stem and loop of yeast tRNA(Phe) (tRNA(ACPhe)) were selected from a random-sequence, 15-amino acid bacteriophage display library. An experimental system, including an affinity selection method, was designed to identify primary RNA-binding peptide sequences without bias to known amino acid sequences and without incorporating nonspecific binding of the anionic RNA backbone. Nitrocellulose binding assays were used to evaluate the binding of RNA by peptide-displaying bacteriophage. Amino acid sequences of RNA-binding bacteriophage were determined from the foreign insert DNA sequences, and peptides corresponding to the RNA-binding bacteriophage inserts were chemically synthesized. Peptide affinities for the RNAs (Kd approximately 0.1-5.0 microM) were analyzed successfully using fluorescence and circular dichroism spectroscopies. These methodologies demonstrate the feasibility of rapidly identifying, isolating, and initiating the analyses of small peptides that bind to RNAs in an effort to define better the chemistry, structure, and function of protein-RNA complexes.
Agris
P F
PF
Department of Biochemistry, North Carolina State University, Raleigh 27695-7622, USA. agris@bch-server.bchncsu.edu
Marchbank
M T
MT
Newman
W
W
Guenther
R
R
Ingram
P
P
Swallow
J
J
Mucha
P
P
Szyk
A
A
Rekowski
P
P
Peletskaya
E
E
Deutscher
S L
SL
D000595:Amino Acid Sequence; D000926:Anticodon; D001435:Bacteriophages; D001483:Base Sequence; D019921:Biotinylation; D002942:Circular Dichroism; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D012342:RNA, Small Nuclear / Q000737:chemistry / Q000378:metabolism*; D012360:RNA, Transfer, Phe / Q000737:chemistry / Q000378:metabolism*; D013050:Spectrometry, Fluorescence
D000926:Anticodon; D019151:Peptide Library; D010455:Peptides; D012342:RNA, Small Nuclear; D012360:RNA, Transfer, Phe