Synthetic, structural and biological studies of the ubiquitin system: the total chemical synthesis of ubiquitin.
The small protein ubiquitin (76 amino acids) has been synthesized under optimized conditions by Merrifield solid-phase methodology using the N alpha-Fmoc protecting group. The crude polypeptide mixture was purified to homogeneity by gel filtration, dialysis and a combination of cation- and anion-exchange chromatography to yield ubiquitin. Amino acid analysis, enzymic digestion and sequencing by automated Edman degradation were used to authenticate the primary structure. Isoelectric focusing and m.s. were used to demonstrate that the final product was greater than 98% pure with a final yield of 93 mg (4.3%) from a single synthesis on a 0.25 nmol scale.
Ramage
R
R
Edinburgh Center for Molecular Recognition, Department of Chemistry, University of Edinburgh, Scotland, U.K.
Green
J
J
Muir
T W
TW
Ogunjobi
O M
OM
Love
S
S
Shaw
K
K
D000595:Amino Acid Sequence; D015153:Blotting, Western; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D007525:Isoelectric Focusing; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D014452:Ubiquitins / Q000138:chemical synthesis* / Q000302:isolation & purification
D014452:Ubiquitins