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Exchanging cofactors in the core antennae from purple bacteria: structure and properties of Zn-bacteriopheophytin-containing LH1.
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The core light-harvesting LH1 complex of Rhodospirillum rubrum consists of an assembly of membrane-spanning alpha and beta polypeptides, each of which binds one bacteriochlorophyll (BChl) a molecule. In this work, we describe a technique that allows the replacement of the natural, Mg BChl a cofactors present in this protein by Zn-bacteriopheophytin (Zn-Bpheo). This technique makes use of the well-characterized, reversible dissociation of LH1 induced by the detergent beta-octylglucoside. Incubation of partially dissociated LH1 with exogeneous pigments induces an equilibrium between the protein-bound BChl and the exogeneous pigment. This results in the binding of chemically modified pigments to LH1, in amounts which depend on the pigment composition and concentration of the exchange buffer. This method can yield information on the relative affinities of the LH1 protein-binding sites for the different pigments BChl and Zn-Bpheo and can also be used to obtain fully reassociated LH1 proteins, with a variable content of modified pigment, which may be precisely monitored. Absorption and FT-Raman spectroscopy indicate that this exchange procedure leads to LH1 proteins containing modified pigments, but retaining a binding site structure identical to that of native LH1. Furthermore, examination of the binding curves suggests that there are two distinguishable binding sites, probably corresponding to the two polypeptides, with very different properties. One of these two binding sites shows a marked preference for Zn-Bpheo over BChl, while the other binding site appears to prefer BChl.
Bacterial Proteins, Bacteriochlorophylls/chemistry, Bacteriochlorophylls/metabolism, Circular Dichroism, Light-Harvesting Protein Complexes, Models, Biological, Models, Chemical, Pheophytins/chemistry, Pheophytins/metabolism, Photosynthetic Reaction Center Complex Proteins/chemistry, Photosynthetic Reaction Center Complex Proteins/metabolism, Pigments, Biological/chemistry, Pigments, Biological/metabolism, Rhodospirillum rubrum/chemistry, Rhodospirillum rubrum/metabolism, Spectrophotometry, Spectrum Analysis, Raman, Structure-Activity Relationship, Zinc/chemistry, Zinc/metabolism
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