Incorporation of beta-selenolo[3,2-b]pyrrolyl-alanine into proteins for phase determination in protein X-ray crystallography

beta -Selenolo[3,2-b]pyrrolyl-L-alanine that mimics tryptophan with the ben zene ring of the indole moiety replaced by selenophene, was incorporated in to human annexin V and barstar. This was achieved by fermentation and expre ssion in a Trp-auxotrophic Escherichia coli host strain using the selective pressure incorporation method. The selenoproteins were obtained in yields comparable to those of the wild-type proteins and exhibit full crystallogra phic isomorphism to the parent proteins, but expectedly show altered absorb ance profiles and quenched tryptophan fluorescence. Since the occurrence of tryptophan residues in proteins is rare, incorporation of the electron-ric h selenium-containing tryptophan surrogate into proteins represents a usefu l supplementation and even a promising novel alternative to selenomethionin e for solving the phase problem in protein X-ray crystallography. (C) 2001 Academic Press.