Cytochrome c maturation: a complex pathway for a simple task?

Post-translational maturation of c-type cytochromes involves covalent attachment of haem to the apocytochrome polypeptide by two thioether bonds. In bacteria, haem attachment occurs in the periplasm, after the separate translocation of haem and the polypeptide across the cytoplasmic membrane. In Escherichia coli, delivery and attachment of the cofactor requires eight or nine specific proteins, which are believed to be organized in a membrane protein complex. After transport across the membrane, haem is attached covalently to the haem chaperone CcmE in an unusual way at a single histidine residue. However, haem binding to CcmE is transient and is succeeded by a further transfer to apocytochrome c. Both haem binding to and release from CcmE involve integral membrane proteins, CcmC and CcmF respectively, which carry a conserved tryptophan-rich motif in a periplasmic domain. Apocytochrome c polypeptides are synthesized as precursors and reach the periplasm by sec-dependent translocation. There they are prepared for haem binding by reduction of the cysteine residues in the motif Cys-Xaa-Xaa-Cys-His, which is characteristic of such proteins. This reduction is achieved in a thio-reduction pathway, whereby electrons are passed from cytoplasmic thioredoxin to the transmembrane protein DsbD, across the membrane, and on to the specific reductases CcmG/CcmH. The merging of the haem delivery and the thio-reduction pathways leads to the stereospecific insertion of haem into various type c cytochromes.     

Marginality,ethnicity and penality: a response to Loïc Wacquant

This article responds to Loïc Wacquant's elaboration of his overarching theoretical framework in his stimulating article ‘Marginality, Ethnicity and Penality in the Neo-liberal City: An Analytic Cartography’. I raise some issues that he might reflect on in his rejoinder to the comments on his essay. These include: (1) the need for greater specification and clarification of explanatory and conceptual variables in the elaboration of his theoretical framework, especially theoretical mechanisms and two key concepts, ‘precariat’ and ‘neo-liberal policy’; and (2) addressing a major study offering empirically based arguments that indirectly challenge assertions in his theoretical framework.     

Cytochrome c assembly: a tale of ever increasing variation and mystery?

Formation of cytochromes c requires a deceptively simple post-translational modification, the formation of two thioether bonds (or rarely one) between the thiol groups of two cysteine residues found in a CXXCH motif (with some occasional variations) and the vinyl groups of heme. There are three partially characterised systems for facilitating this post-translational modification; within these systems there is also variation. In addition, there are clear indications for two other distinct systems. Here some of the current issues in understanding the systems are analysed.     

Multidisciplinary investigation of a ‘British big cat’: a lynx killed in southern England c. 1903

The alleged presence of non-native felid species in the British countryside – popularly, though in part erroneously, known as ‘British big cats’ or ‘alien big cats’ – is a long-standing and controversial topic, perennially of interest to both the mass media and amateur naturalists, and with little apparent acceptance from the technical zoological community. Nevertheless, a number of carcasses and captured live specimens have demonstrated the occasional presence within the region of escapees that potentially explain at least some ‘British big cat’ eyewitness records. We report here the existence of a probable Canada lynx, Lynx canadensis, shot in Newton Abbot, Devon, England, in or prior to 1903, and then accessioned to Bristol Museum and Art Gallery. The specimen (represented by extensive skeletal material and a stuffed taxidermy mount) is Bobcat-like in some respects but is identified as a Canada lynx on the basis of skeletal morphology with a high degree of support; attempts to extract DNA were unsuccessful. Stable strontium isotope analysis supports either a recent introduction from western Canada or long-term acclimation to the local area of Devon where it was collected. Although the specimen was undoubtedly an ‘alien’ (an escapee or release from a collection), it is significant as material evidence in demonstrating, for the first time, the presence of a wild-caught, feral, exotic felid dating to the early years of the twentieth century.     

The novel cytochrome c6 of chloroplasts: a case of evolutionary bricolage?

Cytochrome c6 has long been known as a redox carrier of the thylakoid lumen of cyanobacteria and some eukaryotic algae that can substitute for plastocyanin in electron transfer. Until recently, it was widely accepted that land plants lack a cytochrome c6. However, a homologue of the protein has now been identified in several plant species together with an additional isoform in the green alga Chlamydomonas reinhardtii. This form of the protein, designated cytochrome c6A, differs from the 'conventional' cytochrome c6 in possessing a conserved insertion of 12 amino acids that includes two absolutely conserved cysteine residues. There are conflicting reports of whether cytochrome c6A can substitute for plastocyanin in photosynthetic electron transfer. The evidence for and against this is reviewed and the likely evolutionary history of cytochrome c6A is discussed. It is suggested that it has been converted from a primary role in electron transfer to one in regulation within the chloroplast, and is an example of evolutionary 'bricolage'.     

Long-term c i /c a response of trees in western North America to atmospheric CO2 concentration derived from carbon isotope chronologies

To evaluate how the land carbon reservoir has been responding to the rising CO₂ concentration of the atmosphere, it is important to study how plants in natural forests adjust physiologically to the changing atmospheric conditions. Many experimental studies have addressed this issue, but it has been difficult to scale short-term experimental observations to long-term ecosystem-level responses. This paper derives carbon-isotope-related variables for the past 100–200 years from measurements on trees from natural forests. Calculations show that the c _i/c _a ratios [c _i/c _a is the ratio of the CO₂ concentration (μmol mol⁻¹) in the intercellular space of leaves to that in the atmosphere] of the trees were constant or increased slightly before the 20th century, but changed more rapidly in the 20th century; some increased, some decreased, and some stayed constant. In contrast, the CO₂ concentration inside plant leaves increased monotonically for all trees. Received: 12 June 1997 / Accepted: 29 June 1998     

Primary structure of cytochrome c′ of Methylococcus capsulatus Bath: evidence of a phylogenetic link between P460 and c′-type cytochromes

Cytochrome c′ of Methylococcus capsulatus Bath is involved in electron flow from the enzyme responsible for hydroxylamine oxidation, cytochrome P460, to cytochrome c ₅₅₅. This cytochrome is spectrally similar to other cytochromes c′ but is larger (16,000 Da) and has a lower midpoint potential (–205 mV). By a combination of Edman degradation, mass spectroscopy, and gene sequencing, we have obtained the primary structure of cytochrome c′ from M. capsulatus Bath. The cytochrome shows low sequence similarity to other cytochromes c′, only residues R12, Y53, G56, and the C-terminal heme-binding region (GXXCXXCHXXXK) being conserved. In contrast, cytochrome c′ from M. capsulatus Bath shows considerable sequence similarity to cytochromes P460 from M. capsulatus Bath (31% identity) and from Nitrosomonas europaea (18% identity). This suggests that P460-type cytochromes may have originated from a c′-type cytochrome which developed a covalent cross-link between a lysine residue and the c′-heme. Received: 26 May 1999 / Accepted: 9 September 1999     

The mitochondrial‐derived peptide MOTS‐c: a player in exceptional longevity?

Mitochondrial‐derived peptides (MDP) are encoded by functional short open reading frames in the mitochondrial DNA (mtDNA). These include humanin, and the recently discovered mitochondrial open reading frame of the 12S rRNA‐c (MOTS‐c). Although more research is needed, we suggest that the m.1382A>C polymorphism located in the MOTS‐c encoding mtDNA, which is specific for the Northeast Asian population, may be among the putative biological mechanisms explaining the high longevity of Japanese people.     

Is REP a measure of “objective support”?

Grant and Kluge have recently stated that Bremer support and their own REP (“relative explanatory power”), are the only objective measures of group support. This paper discusses their claim, showing that their philosophical arguments have no basis, and that their own numerical examples actually serve to illustrate shortcomings of REP.     

Extended cardiolipin anchorage to cytochrome c: a model for protein–mitochondrial membrane binding

Two models have been proposed to explain the interaction of cytochrome c with cardiolipin (CL) vesicles. In one case, an acyl chain of the phospholipid accommodates into a hydrophobic channel of the protein located close the Asn52 residue, whereas the alternative model considers the insertion of the acyl chain in the region of the Met80-containing loop. In an attempt to clarify which proposal offers a more appropriate explanation of cytochrome c–CL binding, we have undertaken a spectroscopic and kinetic study of the wild type and the Asn52Ile mutant of iso-1-cytochrome c from yeast to investigate the interaction of cytochrome c with CL vesicles, considered here a model for the CL-containing mitochondrial membrane. Replacement of Asn52, an invariant residue located in a small helix segment of the protein, may provide data useful to gain novel information on which region of cytochrome c is involved in the binding reaction with CL vesicles. In agreement with our recent results revealing that two distinct transitions take place in the cytochrome c–CL binding reaction, data obtained here support a model in which two (instead of one, as considered so far) adjacent acyl chains of the liposome are inserted, one at each of the hydrophobic sites, into the same cytochrome c molecule to form the cytochrome c–CL complex.     

Identification of a novel binding partner of phospholipase cβ1: translin-associated factor X

Mammalian phospholipase Cβ1 (PLCβ1) is activated by the ubiquitous Gα(q) family of G proteins on the surface of the inner leaflet of plasma membrane where it catalyzes the hydrolysis of phosphatidylinositol 4,5 bisphosphate. In general, PLCβ1 is mainly localized on the cytosolic plasma membrane surface, although a substantial fraction is also found in the cytosol and, under some conditions, in the nucleus. The factors that localize PLCβ1in these other compartments are unknown. Here, we identified a novel binding partner, translin-associated factor X (TRAX). TRAX is a cytosolic protein that can transit into the nucleus. In purified form, PLCβ1 binds strongly to TRAX with an affinity that is only ten-fold weaker than its affinity for its functional partner, Gα(q). In solution, TRAX has little effect on the membrane association or the catalytic activity of PLCβ1. However, TRAX directly competes with Gα(q) for PLCβ1 binding, and excess TRAX reverses Gα(q) activation of PLCβ1. In C6 glia cells, endogenous PLCβ1 and TRAX colocalize in the cytosol and the nucleus, but not on the plasma membrane where TRAX is absent. In Neuro2A cells expressing enhanced yellow and cyano fluorescent proteins (i.e., eYFP- PLCβ1 and eCFP-TRAX), F?rster resonance energy transfer (FRET) is observed mostly in the cytosol and a small amount is seen in the nucleus. FRET does not occur at the plasma membrane where TRAX is not found. Our studies show that TRAX, localized in the cytosol and nucleus, competes with plasma-membrane bound Gα(q) for PLCβ1 binding thus stabilizing PLCβ1 in other cellular compartments.     

反相HPLC法测定四种柴胡的柴胡皂式a,c,d含量

1990版《中国药典》规定正品柴胡为北柴胡和红柴胡,而各地实际入药的柴胡种类往往与药典规定不尽相同。因此有必要对目前已供药用的柴胡生药中有效成分进行定量分析,作为评判各种柴胡质量的依据之一。本文应用反相HPLC法测定了9种市售商品柴胡,经鉴定为北柴胡（BupleurumchinenseDC．）、红柴胡（B．scorzonerifoliumWilld．）、空心柴胡（B．longicauleWall．exDC．var．franchetiideBoiss．）和不同产地的小叶黑柴胡（B．smithiiWolffvar．parvifoliumShanetY．Li）,分别测定了其柴胡皂式a,c,d（简记为ssa,ssc,ssd）的含…     

The mycoflora and toxicity of feedstuffs from a production plant in córdoba, Argentina

Feedstuffs used for poultry nutrition in Argentina were analyzed for fungal flora and natural incidence of mycotoxins. Survey of 120 samples of poultry feeds, taken from May 1998 to April 1999, showed the presence of 15 genera of filamentous fungi. The predominant genera wereFusarium spp. andPenicillium ssp., isolated in 67.5 % of the samples, followed byAspergillus spp. (57.5 %). Yeast, were significantly isolated from most of the samples. Species identification was carried down for the toxigenic genera. Fungal total counts of poultry feeds ranged from 2.0 × 10³ to 3.0 × 10⁵ CFU g^-1 The fungal total counts during two months of sampling, were slightly over the limit value of 1 × 105 CFU g^-1, which ensure the hygienic quality of the feed. Potentially toxicogenic species presented moderate mean colony counts. Many of the fungi isolated from poultry feeds are mycotoxin producers. Fumonisins had the highest incidence, and were found in 97 % of the analyzed samples followed by aflatoxin B₁ (46 %), zearalenone (18 %) and deoxynivalenol (6 %). On the co-occurrence of both carcinogenic mycotoxins, all of the FBs contaminated feed samples were co-contaminated with AFB₁. The results show the relevance of the samples screening for viable fungi propagules and the surveillance of their associated mycotoxins in poultry feeds.     

Stabilization of mesophilic Allochromatium vinosum cytochrome c′ through specific mutations modeled by a thermophilic homologue

AVCP cytochrome c′ from mesophilic Allochromatium vinosum exhibits lower stability than a thermophilic counterpart, Hydrogenophilus thermoluteolus cytochrome c′ (PHCP), in which the six specific amino acid residues that are not conserved in AVCP are responsible for its stability. Here we measured the stability of AVCP variants carrying these specific residues instead of the original AVCP ones. Among the six single AVCP variants, all of which formed a dimeric structure similar to that of the wild-type, three were successfully stabilized compared with the wild-type, while one showed lower stability than the wild-type. In addition, the most stabilized and destabilized AVCP variants could bind CO, similar to the wild-type. These results indicated that mesophilic AVCP could be stabilized through specific three mutations modeled by the thermophilic counterpart, PHCP, without changing the CO binding ability.     

Ab initio structure solution of a dimeric cytochrome c 3 from Desulfovibrio gigas containing disulfide bridges

The 1.2?Å resolution crystal structure of the 29?kDa di-tetrahaem cytochrome c ₃ from the sulfate reducing bacterium Desulfovibrio gigas was solved by ab initio methods, making this the largest molecule to be solved by this procedure. The actual refined model of the cysteine-linked dimeric molecule reveals that this molecule is very similar to the non-covalently linked symmetrical dimer of the di-tetrahaem cytochrome c ₃ from Desulfomicrobium norvegicum. Each monomer has the typical polypeptide fold, haem arrangement and iron coordination found for the tetrahaem cytochrome c ₃ molecules. The interface between the covalently linked monomers in the asymmetric unit of the crystal shows a pseudo two-fold arrangement, disturbed from symmetry by crystal packing forces. The fact that D. gigas contains a dimeric tetrahaem cytochrome with solvent accessible disulfide bridges and that this cytochrome specifically couples hydrogen oxidation to thiosulfate reduction in bacterial extracts provides an interesting aspect related to disulfide exchange reactions in this microorganism.