Consequences for the Organization of Reaction Center-Light Harvesting Antenna 1 (LH1) Core Complexes of Rhodobacter sphaeroides Arising from Deletion of Amino Acid Residues from the C Terminus of the LH1 α Polypeptide

Abstract

The light harvesting antenna 1 (LH1) complex of Rhodobacter sphaeroides is intimately associated with the reaction center (RC) as part of the reaction center RC-LH1 core complex. The pufA gene has been modified such that between 5 and 16 amino acid residues were progressively deleted from the C terminus of the LH1 ␣ polypeptide. The two largest deletions produced strains which were deficient in LH1. The remaining four deletion mutants exhibited significant reductions in the average level of LH1 per reaction center. Analysis of detergent-solubilized cores on sucrose gradients showed that the mutant strains had a sizeable population of antenna-deficient reaction centers in addition to core complexes with a reduced ratio of LH1:RC. The decrease in the ratio of LH1:RC in core complexes of the mutant strains was accompanied by a progressive blue shift of the absorbance maximum of LH1, which we attribute to the reduced aggregation state of LH1 in the smaller cores. The PufX polypeptide was not required for photosynthetic growth in mutants with reduced core sizes. We conclude that the level of LH1 in the bacterial membrane, and the aggregation state of LH1 in core complexes, are both influenced by the C terminus of the ␣ polypeptide, and we discuss possible models for the organization of the core complex in Rb. sphaeroides.The photosynthetic bacterium Rhodobacter sphaeroides has a relatively simple membrane-bound photosystem comprising a single type of reaction center (RC) 1 and two types of light harvesting complex. The peripheral light harvesting complex, LH2, is present in variable amounts according to the incident light intensity whereas the core light harvesting complex, LH1, is present in a fixed stoichiometry to the RC (1, 2). In the simple "lake" model (3) light energy captured by LH2 migrates via LH1 to a dimer of bacteriochlorophylls (Bchl) within the RC, initiating the transfer of an electron through the RC from the periplasmic side to the cytoplasmic side of the membrane. The products of this transmembrane electron transfer, reduced ubiquinone and an oxidized Bchl dimer, trigger a cycle of electron transfer reactions involving the intramembrane pool of ubiquinone, the cytochrome bc 1 complex and a soluble cytochrome c that results in the translocation of protons from the cytoplasmic side of the membrane to the periplasmic side, generating a proton electrochemical gradient.All bacterial light harvesting complexes studied to date contain two small hydrophobic polypeptides designated ␣ and ␤ in a 1:1 ratio, both of which have a single membrane-spanning helix (4). Liganded to these polypeptides are the light harvesting pigments which, in the case of Rb. sphaeroides, are molecules of Bchl a. Carotenoids also act as light harvesting pigments but have an additional photoprotective role. In the LH2 complex of Rb. sphaeroides each pair of ␣ and ␤ subunits is associated with three molecules of Bchl, two of which absorb at 850 nm and the third at 800 nm. Within the LH1 complex there are only two Bchl molecules per pai...

The long-range supraorganization of the bacterial photosynthetic unit: A key role for PufX

“…sphaeroides (6), and more specifically that it is required for ubiquinone͞ubiquinol exchange between the RC Q B site and the cytochrome bc 1 complex (7,8). When the LH1 complex is absent or is reduced in size, there is no such requirement for PufX (9,10). Thus, there is an intriguing relationship between the RC, the LH1, and cytochrome bc 1 complex, in which PufX plays a central role.…”

“…The LH1 complex forms a ring apparently consisting of 16 ␣, ␤ heterodimers (4), which then surrounds the RC, at least in the absence of PufX (9,12,10). Although there are no high resolution data for any LH1 complex, it is assumed that this complex contains a maximum of 32 BChls sandwiched between concentric rings of ␣ and ␤ subunit helices (4), much like arrangement found in the LH2 complex (3).…”

“…Although there are no high resolution data for any LH1 complex, it is assumed that this complex contains a maximum of 32 BChls sandwiched between concentric rings of ␣ and ␤ subunit helices (4), much like arrangement found in the LH2 complex (3). In the presence of PufX, the number of BChls in the ring is reduced by approximately 2 (10). PufX is believed to form part of the LH1 ring (12); it associates with LH1 and the RC in vitro to form the core complex in Rb.…”

The purple bacterial photosynthetic unit

“…In this case, a small increase in the ratio of LH1 Bchls a:RC is observed in comparison with the model in (A) (McGlynn et al 1994(McGlynn et al , 1996Barz et al 1995a et al 1990, 1992) using reaction center only and reaction center-LH1 strains, McGlynn et al (1994) were able to show that the loss of Puf X was deleterious only when the LH1 was present. In a further development of this approach, strains were constructed in which the LHI: reaction center 'core' size was genetically modified (McGlynn et al 1996). This manipulation resulted in a progressive truncation of the C-terminal region of the LH1 c~ apoprotein.…”

Overexpression of pucC improves the heterologous protein expression level in a Rhodobacter sphaeroides expression system

“…The LH2 complex has typical absorption at ~800 and ~850 nm, which can be used for spectral detection of the expression of LH2 as well as the presence of LH2 as a β-subunit in fusion proteins (Golecki et al, 1979;McGlynn et al, 1996;Hu et al, 2002). Thus, in previous studies, we have developed a rapid heterologous protein evaluation method in a R. sphaeroides expression system according to its spectral characteristics (Zhao et al, 2011).…”