Structure and function of the photosynthetic reaction center fromRhodobacter sphaeroides

The three-dimensional structure of the photosynthetic reaction center fromRhodobacter sphaeroides is described. The reaction center is a transmembrane protein that converts light into chemical energy. The protein has three subunits: L, M, and H. The mostly helical L and M subunits provide the scaffolding and the finely tuned environment in which the chromophores carry out electron transfer. The details of the protein-chromophore interactions are from studies of a trigonal crystal form that diffracted to 2.65-Å resolution. Functional studies of the multi-subunit complex by site-specific replacement of key amino acid residues are summarized in the context of the molecular structure.This work was supported in part by the U.S. Department of Energy, Office of Health and Environmental Research, under Contract No. W-31-109-ENG-38 and by Public Health Service Grant GM36598.     

Effect of exchangeable aluminium on the reduction of potato scab

The severity of the incidence of the fungal disease, potato scab, varies with different soil groups at the same soil pH. At a soil pH of 5.3, potato scab is easily controlled in soils of western Hokkaido (soil group A) by simply decreasing soil pH, but in soils from eastern Hokkaido (soil group B) it is not so easily controlled. The difference appears to be due to higher levels and exchangeable aluminium in Group A.Addition of sufficient aluminium or ferrous sulfate to a group B soil decreased the incidence of potato scab in a field experiment. Higher levels of aluminium sulfate depressed crop production. It is concluded that aluminium ions control the incidence of potato scab in acid soils. It is suggested that, in soils with low exchange acidity Y1, potato scab can be controlled by adding sufficient aluminium to increase their exchange acidity Y1 to above 7–8.     

Nonlinear effect of dynamic self-organization in macromolecular systems caused by photocontrolled electron flux

We use the electron-conformational interaction approach to develop a physical model which self-consistently describes the photomobilized electron transfer kinetics and structure conformational transitions in reaction centers (RCs) of purple bacteria. We consider the kinetics of electron transition from pigment onto primary acceptor and the subsequent charge recombination accounting for the change of distance between the above-mentioned cofactors. It is shown that, given natural values of RC parameters, the kinetic constant's dependence on the acting light intensity is monotone. As opposed to the previous case, similar dependencies for the chain of electron transfer between primary and secondary quinone acceptors revealed anS-like relationship. This can lead to bistability of the RC optical transmission coefficient and a fundamental dependence of charge recombination kinetics upon the prehistory of the RC's interaction with exciting radiation.     

Possible mechanism of nitric oxide production from N-hydroxy-l-arginine or hydroxylamine by superoxide ion

It has been speculated that N^G-hydroxy-l-arginine (OH-l-Arg), which is an intermediate in NO production from l-arginine, may be converted to NO by superoxide ion. However, there is still no direct evidence for this conversion. In the present study this was investigated using superoxide ion generated either in acellular or cellular systems. It was found that OH-l-Arg and hydroxylamine were converted to nitrite and nitrate apparently via NO by superoxide ion in aqueous solution. Arginine remained unaffected. These changes were observed during reaction of chemical substances as well as in a biological system (zymosan-activated macrophages in culture). Superoxide dismutase prevented this transformation. OH-l-Arg was also spontaneously hydrolysed to hydroxylamine and l-citrulline, however this occurred at pH > 9 only. Activated microsomes (containing different isoforms of cytochrome P450) were unable to replace NO-synthase in its ability to produce OH-l-Arg from l-arginine. These data support the hypothesis that a pathway alternative to the well-known synthesis of NO by NO-synthase via OH-l-Arg exists. This pathway may involve the production of OH-l-Arg by NO-synthase and decomposition of OH-l-Arg to NO by the action of superoxide ion. Alternatively, hydrolysis of OH-l-Arg to hydroxylamine may occur followed by its oxidation to NO, again by superoxide ion.     

Inhibition by trifluoperazine of ATP synthesis and hydrolysis by particulate and soluble mitochondrial F1: Competition with H2PO 4 −

The effect of trifluoperazine (TFP) on the ATPase activity of soluble and paniculate F₁ATPase and on ATP synthesis driven by succinate oxidation in submitochondrial particles from bovine heart was studied at pH 7.4 and 8.8. At the two pH. TFP inhibited ATP hydrolysis. Inorganic phosphate protected against the inhibiting action of TFP. The results on the effect of various concentrations of phosphate in the reversal of the action of TFP on hydrolysis at pH 7.4 and 8.8 showed that H₂PO ₄ ^– is the species that competes with TFP. The effect of TFP on oxidative phosphorylation was studied at concentrations that do not produce uncoupling or affect the aerobic oxidation of succinate (<15M). TFP inhibited oxidative phosphorylation to a higher extent at pH 8.8 than at pH 7.4; this was through a diminution in theV _max, and an increase in theK _m for phosphate. Data on phosphate uptake during oxidative phosphorylation at several pH showed that H₂PO ₄ ^– is the true substrate for oxidative phosphorylation. Thus, in both synthesis and hydrolysis of ATP, TFP and H₂PO ₄ ^– interact with a common site. However, there is a difference in the sensitivity to TFP of ATP synthesis and hydrolysis; this is more noticeable at pH 8.8, i.e. ATPase activity of soluble F₁ remains at about 40% of the activity of the control in a concentration range of TFP of 40–100M, whereas in oxidative phosphorylation 14M TFP produces a 60% inhibition of phosphate uptake.     

Analysis of the proposed Fe-SX binding region of Photosystem 1 by site directed mutation of PsaA in Chlamydomonas reinhardtii

The psaA and psaB genes of the chloroplast genome in oxygenic photosynthetic organisms code for the major peptides of the Photosystem 1 reaction center. A heterodimer of the two polypeptides PsaA and PsaB is thought to bind the reaction center chlorophyll, P700, and the early electron acceptors A₀, A₁ and Fe-S_X. Fe-S_X is a 4Fe4S center requiring 4 cysteine residues as ligands from the protein. As PsaA and PsaB have only three and two conserved cysteine residues respectively, it has been proposed by several groups that Fe-S_X is an unusual inter-peptide center liganded by two cysteines from each peptide. This hypothesis has been tested by site directed mutagenesis of PsaA residue C575 and the adjacent D576. The C575D mutant does not assemble Photosystem 1. The C575H mutant contains a photoxidisable chlorophyll with EPR properties of P700, but no other Photosystem 1 function has been detected. The D576L mutant assembles a modified Photosystem 1 in which the EPR properties of the Fe-S_A/B centers are altered. The results confirm the importance of the conserved cysteine motif region in Photosystem 1 structure.Dedicated to the memory of Daniel I. Arnon.     

Sodium channels in cultured neuroblastoma cells grown in high glucose or l-Fucose

Patch clamp techniques were used to record whole cell and single channel Na⁺ currents from NB41A3 neuroblastoma cells grown in culture. Cells were grown for two weeks in control medium or medium supplemented with 30 mm d-glucose of 30 mm l-fucose.Cells exposed to glucose or l-fucose had smaller whole cell Na⁺ currents than cells grown in unsupplemented medium, consistent with earlier studies (Yorek, Stefani & Wachtel, 1994). Whole cell macroscopic currents showed no change in activation or inactivation kinetics. Single channel current properties and opening probability were also unchanged.The number of [³H]saxitoxin binding sites, and therefore the total number of Na⁺ channels, was not reduced in cells grown in glucose or l-fucose (Yorek et al., 1994). Therefore, we conclude that some of the channels must have been rendered nonfunctional by the conditioning media. The finding that single channel properties are not altered suggests that channels become nonfunctional in an all-or-none manner.This work was supported by Merit Review Awards to M.A. Yorek and R.E. Wachtel from the Department of Veterans Affairs and by National Institutes of Health grant DK45453 to M.A. Yorek.     

The UDP-Galactose:Ceramide Galactosyltransferase: Expression Pattern in Oligodendrocytes and Schwann Cells During Myelination and Substrate Preference for Hydroxyceramide

Abstract: Galactosylceramide ("galactocerebroside"; GalC) is a major glycolipid in the myelin sheath of the CNS and the PNS. The enzyme UDP-galactose:ceramide galactosyltransferase (CGalT) catalyzes the final step of the synthesis of GalC: the transfer of galactose to ceramide. By a differential screening approach, we have isolated a cDNA, the sequence of which is identical to the recently isolated cDNA clones for CGalT. By northern analysis and in situ hybridization we demonstrated that CGalT mRNA is expressed at birth in oligodendrocytes and Schwann cells, an expression pattern corresponding to the onset of myelination. In addition to the high expression levels of CGalT in oligodendrocytes and Schwann cells, in situ hybridization also showed expression in subtypes of neurons in spinal cord, cerebellum, and brainstem in the adult CNS, but at a much lower level than in oligodendrocytes. Expression of CGalT in COS cells demonstrated that CGalT has a preference for hydroxyceramide as a substrate. CGalT-expressing COS cells synthesize and transport GalC to their cell surface as shown by immunofluorescence and by lipid analysis of living cells. Our results suggested that the CGalT specifically uses hydroxyceramide for the synthesis of GalC and that separate (co)enzymes are not needed.