Retinal migration during dark reduction of bacteriorhodopsin.
When the retinal Schiff base in chymotryptically cleaved bacteriorhodopsin is reduced to a secondary retinylamine by prolonged exposure to 10% (wt/vol) sodium cyanoborohydride, at pH 10, in the absence of light, approximately 45% of the retinal is found linked to Lys-41 and 22% to Lys-40, and the remainder is scattered over various sites on the large chymotryptic fragment, including the physiological site at Lys-216.  The retinal-binding site is destroyed or blocked by the reduction conditions, but the bacteriorhodopsin lattice remains intact.  The results demonstrate that artifactual linkage to Lys-40/41 is possible under special conditions.  Under these conditions, the epsilon-amino groups of Lys-40/41 show an enhanced ability to form retinylidene linkages with the retinal released by the physiological linkage site at Lys-216, due to some combination of close proximity to the normal linkage site, and increased reactivity with respect to other lysine epsilon-amino groups.  The results are of interest for the characterization of the two newly discovered rhodopsin-like proteins, halorhodopsin and slow rhodopsin.
Wolber
P K
PK
Department of Biochemistry and Biophysics, University of California, San Francisco 94143, USA.
Stoeckenius
W
W
D000596:Amino Acids; D001436:Bacteriorhodopsins / Q000737:chemistry*; D001894:Borohydrides; D005591:Chemical Fractionation; D002918:Chymotrypsin / Q000737:chemistry; D003488:Cyanogen Bromide / Q000737:chemistry; D003624:Darkness; D019617:Halobacterium salinarum / Q000502:physiology*; D006863:Hydrogen-Ion Concentration; D008027:Light; D011107:Polylysine / Q000378:metabolism; D011485:Protein Binding; D018717:Purple Membrane / Q000737:chemistry*; D012172:Retinaldehyde / Q000031:analogs & derivatives* / Q000737:chemistry*; D012545:Schiff Bases; D014316:Tritium
D000596:Amino Acids; D001894:Borohydrides; D012545:Schiff Bases; D014316:Tritium; D011107:Polylysine; D001436:Bacteriorhodopsins; C009282:sodium cyanoborohydride; D002918:Chymotrypsin; D003488:Cyanogen Bromide; D012172:Retinaldehyde