Location of Chemically Modified Lysine 41 in the Structure of Bacteriorhodopsin by Neutron Diffraction

• Published in: Biophysical journal Vol. 50; no. 4; pp. 629 - 635
• Main Authors: Seiff, F., Wallat, I., Westerhausen, J., Heyn, M.P.
• Format: Journal Article
• Language: English
• Published: Bethesda, MD Elsevier Inc 01.10.1986; Biophysical Society
• Subjects: Applied sciences; Biological and medical sciences; Cell structures and functions; Chloroplast, photosynthetic membrane and photosynthesis; Exact sciences and technology; Fundamental and applied biological sciences. Psychology; Molecular and cellular biology; Other techniques and industries; Bacteriorhodopsin; Site specificity; Chemical modification; Photosystem; Archaeobacteria; Neutron diffraction; Lysine; Purple membrane; Bacteria; Halobacteriaceae; Localization; Halobacterium halobium
• ISSN: 0006-3495; 1542-0086
• DOI: 10.1016/S0006-3495(86)83502-0

Purple membranes were prepared in which bacteriorhodopsin was labeled at lysine 41 with phenylisothiocyanate (PITC) and with perdeuterated PITC. The in-plane position of this small label containing only five deuterons was determined from the differences between the neutron diffraction intensities of the two samples. At 8.7-Å resolution the Fourier difference map revealed a well-defined site between helices 3 and 4. This position was confirmed by a refinement procedure in reciprocal space. Model calculations showed that the observed difference density had the right amplitude for the label. Thus it is possible to locate a small group in a large protein structure by replacing as few as five hydrogens by deuterium. The observed location of PITC restricts the number of possibilities for the assignment of helix B in the sequence (to which lysine 41 is attached) to one of the seven helices of the structure. Taking into account the size of the label and the length of the lysine side chain our result excludes helices 1, 2, and 7 as candidates for B.