Structural role of (bacterio)chlorophyll ligated in the energetically unfavorable beta-position

Chlorophyll is attached to apoprotein in diastereotopically distinct ways, by beta- and alpha-ligation. Both the beta- and alpha-ligated chlorophylls of photosystem I are shown to have ample contacts to apoprotein within their proteinaceous binding sites, in particular, at C-13 of the isocyclic ring. The H-bonding patterns for the C-13(1) oxo groups, however, are clearly distinct for the beta-ligated and alpha-ligated chlorophylls. The beta-ligated chlorophylls frequently employ their C-13(1) oxo in H-bonds to neighboring helices and subunits. In contrast, the C-13(1) oxo of alpha-ligated chlorophylls are significantly less involved in H-bonding interactions, particularly to neighboring helices. Remarkably, in the peripheral antenna, light harvesting complex (LH2) from Rhodobacter sphaeroides, a single mutation in the alpha-subunit, introduced to eliminate H-bonding to the beta-bacteriochlorophyll-B850, which is ligated in the "beta-position," results in significant thermal destabilization of the LH2 in the membrane. In addition, in comparison with wild type LH2, the expression level of the LH2 lacking this H-bond is significantly reduced. These findings show that H-bonding to the C-13(1) keto group ofbeta-ligated (bacterio)-chlorophyll is a key structural motif and significantly contributes to the stability of bacteriochlorophyll proteins in the native membrane. Our analysis of photosystem I and II suggests that this hitherto unrecognized motif involving H-bonding to beta-ligated chlorophylls may be equally critical for the stable assembly of the inner core antenna of these multicomponent chlorophyll proteins.