Biomedical subjects

J Castresana

Publications and source records attributed to J Castresana.

At least 19 recordsLinked to original sources

PubMed2001

Comparative genomics and bioenergetics.

Bacterial and archaeal complete genome sequences have been obtained from a wide range of evolutionary lines, which allows some general conclusions about the phylogenetic distribution and evolution of bioenergetic pathways to be drawn. In particular, I searched in the complete genomes for key enzymes involved in aerobic and anaerobic respiratory pathways and in photosynthesis, and mapped them into an rRNA tree of sequenced species. The phylogenetic distribution of these enzymes is very irregular, and clearly shows the diverse strategies of energy conservation used by prokaryotes. In addition, a thorough phylogenetic analysis of other bioenergetic protein families of wide distribution reveals a complex evolutionary history for the respective genes. A parsimonious explanation for these complex phylogenetic patterns and for the irregular distribution of metabolic pathways is that the last common ancestor of Bacteria and Archaea contained several members of every gene family as a consequence of previous gene or genome duplications, while different patterns of gene loss occurred during the evolution of every gene family. This would imply that the last universal ancestor was a bioenergetically sophisticated organism. Finally, important steps that occurred during the evolution of energetic machineries, such as the early evolution of aerobic respiration and the acquisition of eukaryotic mitochondria from a proteobacterium ancestor, are supported by the analysis of the complete genome sequences.

Archaea↗

PubMed2001

A molecular phylogeny of two extinct sloths.

Xenarthra (Edentata) is an extremely diverse mammalian order whose modern representatives are the armadillos, anteaters, and sloths. The phylogeny of these groups is poorly resolved. This is particularly true for the sloths (phyllophagans), originally a large and diverse group now reduced to two genera in two different families. Both morphological analyses and molecular analyses of rDNA genes of living and extinct sloths have been used with limited success to elucidate their phylogeny. In an attempt to clarify relationships among the sloths, DNA was extracted and mitochondrial cytochrome b gene sequences were determined from representatives of two extinct groups of sloths (Mylodontidae and Megatheriidae), their two living relatives (two-toed sloths [Megalonychidae], three-toed sloths [Bradypodidae]), anteaters and armadillos. A consistent feature of the latter two species was the nuclear copies of cytochrome b gene sequences. Several methods of phylogenetic reconstruction were applied to the sequences determined, and the results were compared with 12S rDNA sequences obtained in previous studies. The cytochrome b gene exhibited a phylogenetic resolving power similar to that of the 12S rDNA sequences. When both data sets were combined, they tended to support the grouping of two-toed sloths with mylodontids and three-toed sloths with megatheriids. The results strengthen the view that the two families of living sloths adapted independently to an arboreal life-style.

Animals↗

PubMed2001

Cytochrome b phylogeny and the taxonomy of great apes and mammals.

In the Linnaean system of classification, the generic status of a species is part of its binomial name, and it is therefore important that the classification at the level of genus is consistent at least in related groups of organisms. Using maximum-likelihood phylogenetic trees constructed from a large number of complete or nearly complete mammalian cytochrome b sequences, I show that the distributions of intrageneric and intergeneric distances derived from these trees are clearly separated, which allows the limits for a more rational generic classification of mammals to be established. The analysis of genetic distances among hominids in this context provides strong support for the inclusion of humans and chimpanzees in the same genus. It is also of interest to decipher the main reasons for the possible biases in the mammalian classification. I found by correlation analysis that the classification of mammals of large body size tends to be oversplit, whereas that of small mammals has an excess of lumping, which may be a manifestation of the larger difficulty in finding diagnostic characters in the classification of small animals. In addition, and contrary to some previous observations, there is no correlation between body size and rate of cytochrome b evolution in mammals, which excludes the difference in evolutionary rates as the cause of the observed body size taxonomic bias.

Animals↗

PubMed2000

Nitric oxide reductases in bacteria.

Nitric oxide reductases (NORs) that are found in bacteria belong to the large enzyme family which includes cytochrome oxidases. Two types of bacterial NORs have been characterised. One is a cytochrome bc-type complex (cNOR) that receives electrons from soluble redox protein donors, whereas the other type (qNOR) lacks the cytochrome c component and uses quinol as the electron donor. The latter enzyme is present in several pathogens that are not denitrifiers. We summarise the current knowledge on bacterial NORs, and discuss the evolutionary relationship between them and cytochrome oxidases in this review.

Amino Acid Sequence↗

PubMed2000

Selection of conserved blocks from multiple alignments for their use in phylogenetic analysis.

The use of some multiple-sequence alignments in phylogenetic analysis, particularly those that are not very well conserved, requires the elimination of poorly aligned positions and divergent regions, since they may not be homologous or may have been saturated by multiple substitutions. A computerized method that eliminates such positions and at the same time tries to minimize the loss of informative sites is presented here. The method is based on the selection of blocks of positions that fulfill a simple set of requirements with respect to the number of contiguous conserved positions, lack of gaps, and high conservation of flanking positions, making the final alignment more suitable for phylogenetic analysis. To illustrate the efficiency of this method, alignments of 10 mitochondrial proteins from several completely sequenced mitochondrial genomes belonging to diverse eukaryotes were used as examples. The percentages of removed positions were higher in the most divergent alignments. After removing divergent segments, the amino acid composition of the different sequences was more uniform, and pairwise distances became much smaller. Phylogenetic trees show that topologies can be different after removing conserved blocks, particularly when there are several poorly resolved nodes. Strong support was found for the grouping of animals and fungi but not for the position of more basal eukaryotes. The use of a computerized method such as the one presented here reduces to a certain extent the necessity of manually editing multiple alignments, makes the automation of phylogenetic analysis of large data sets feasible, and facilitates the reproduction of the final alignment by other researchers.

Amino Acid Sequence↗

PubMed1999

Respiratory chains in the last common ancestor of living organisms.

Sequences in current databases show that a number of proteins involved in respiratory processes are homologous in archaeal and bacterial species. In particular, terminal oxidases belonging to oxygen, nitrate, sulfate, and sulfur respiratory pathways have been sequenced in members of both domains. They include cytochrome oxidase, nitrate reductase, adenylylsulfate reductase, sulfite reductase, and polysulfide reductase. These proteins can be assigned to the last common ancestor of living organisms assuming that the deepest split of the three domains of life occurred between Archaea and Bacteria and that they were not acquired through lateral gene transfer by one of these domains. These molecular data indicate that several of the most important respiratory pathways arose early in evolution and that the last common ancestor of living organisms was not a simple organism in its energetic metabolism. Rather, it may have been able to gain energy by means of at least four electron transport chains, and therefore it may have been prepared to face a wide range of environmental conditions.

Archaea↗

PubMed1998

Codon reassignment and amino acid composition in hemichordate mitochondria.

In the mitochondrial genome of the hemichordate Balanoglossus carnosus, the codon AAA, which is assigned to lysine in most metazoans but to asparagine in echinoderms, is absent. Furthermore, the lysine tRNA gene carries an anticodon substitution that renders its gene product unable to decode AAA codons, whereas the asparagine tRNA gene has not changed to encode a tRNA with the ability to recognize AAA codons. Thus, the hemichordate mitochondrial genome can be regarded as an intermediate in the process of reassignment of mitochondrial AAA codons, where most metazoans represent the ancestral situation and the echinoderms the derived situation. This lends support to the codon capture hypothesis. We also show that the reassignment of the AAA codon is associated with a reduction in the relative abundance of lysine residues in mitochondrial proteins.

Amino Acids↗

PubMed1998

The mitochondrial genome of the hemichordate Balanoglossus carnosus and the evolution of deuterostome mitochondria.

The complete nucleotide sequence of the mitochondrial genome of the hemichordate Balanoglossus carnosus (acorn worm) was determined. The arrangement of the genes encoding 13 protein, 22 tRNA, and 2 rRNA genes is essentially the same as in vertebrates, indicating that the vertebrate and hemichordate mitochondrial gene arrangement is close to that of their common ancestor, and, thus, that it has been conserved for more than 600 million years, whereas that of echinoderms has been rearranged extensively. The genetic code of hemichordate mitochondria is similar to that of echinoderms in that ATA encodes isoleucine and AGA serine, whereas the codons AAA and AGG, whose amino acid assignments also differ between echinoderms and vertebrates, are absent from the B. carnosus mitochondrial genome. There are three noncoding regions of length 277, 41, and 32 bp: the larger one is likely to be equivalent to the control region of other deuterostomes, while the two others may contain transcriptional promoters for genes encoded on the minor coding strand. Phylogenetic trees estimated from the inferred protein sequences indicate that hemichordates are a sister group of echinoderms.

Animals↗

PubMed1998

A hominoid-specific nuclear insertion of the mitochondrial D-loop: implications for reconstructing ancestral mitochondrial sequences.

A nuclear integration of a mitochondrial control region sequence on human chromosome 9 has been isolated. PCR analyses with primers specific for the respective insertion-flanking nuclear regions showed that the insertion took place on the lineage leading to Hominoidea (gibbon, orangutan, gorilla, chimpanzee, and human) after the Old World monkey-Hominoidea split. The sequences of the control region integrations were determined for humans, chimpanzees, gorillas, orangutans, and siamangs. These sequences were then used to construct phylogenetic trees with different methods, relating them with several hominoid, Old Work monkey, and New World monkey mitochondrial control region sequences. Applying maximum-likelihood, neighbor-joining, and parsimony algorithms, the insertion clade was attached to the branch leading to the hominoid mitochondrial sequences as expected from the PCR-determined presence/absence of this integration. An unexpected long branch leading to the internal node that connects all insertion sequences was observed for the different phylogeny reconstruction procedures. This finding is not totally compatible with the lower evolutionary rate in the nucleus than in the mitochondrial compartment. We determined the unambiguous substitutions on the branch leading to the most recent common ancestor (MRCA) of the mitochondrial inserts according to the parsimony criterium. We propose that they are unlikely to have been caused by damage of the transposing nucleic acid and that they are probably due to a change in the evolutionary mode after the transposition.

Animals↗

PubMed1997

Evolution of the spectrin repeat.

We now know that the evolution of multidomain proteins has frequently involved genetic duplication events. These, however, are sometimes difficult to trace because of low sequence similarity between duplicated segments. Spectrin, the major component of the membrane skeleton that provides elasticity to the cell, contains tandemly repeated sequences of 106 amino acid residues. The same repeats are also present in alpha-actinin, dystrophin and utrophin. Sequence alignments and phylogenetic trees of these domains allow us to interpret the evolutionary relationship between these proteins, concluding that spectrin evolved from alpha-actinin by an elongation process that included two duplications of a block of seven repeats. This analysis shows how a modular protein unit can be used in the evolution of large cytoskeletal structures.

Amino Acid Sequence↗

PubMed1995

Does Vav bind to F-actin through a CH domain?

An actin-binding protein domain we call here 'calponin-homology' or CH is present in signalling proteins such as Vav which are involved in activation and inactivation of small G-proteins. Using profile methods, we have detected two repeats of this domain in the actin-binding region of alpha-actinin and related proteins. Based on this, we propose that CH domain in Vav and other signalling proteins is employed for association with filamentous actin, and that this function correlates with their control on the G-proteins Rac and Rho which are involved in the organization of cytoskeleton.

Actins↗

PubMed1995

New archaebacterial genes coding for redox proteins: implications for the evolution of aerobic metabolism.

Archaebacterial respiratory chains are poorly understood at the molecular level. We have cloned and sequenced a cluster of five new genes from the archaebacterium Sulfolobus acidocaldarius, four of them coding for redox proteins: a Rieske iron-sulphur protein, a cytochrome b, a subunit II of cytochrome oxidase and a blue copper protein (sulfocyanin). The fifth gene codes for a hydrophobic protein with no homologue in the databases. The gene organization and biochemical data suggest that all four redox proteins probably form part of a membrane respiratory complex together with SoxM, a previously characterized catalytic subunit of cytochrome oxidase. A phylogenetic analysis of the new protein sequences gives support to the view that an elaborate aerobic respiratory chain was already present in the last common ancestor of all living organisms.

Aerobiosis↗

PubMed1995

Evolution of energetic metabolism: the respiration-early hypothesis.

The main energy-transducing metabolic systems originated and diversified very early in the evolution of life. This makes it difficult to unravel the precise steps in the evolution of the proteins involved in these processes. Recent molecular data suggest that homologous proteins of aerobic respiratory chains can be found in Bacteria and Archaea, which points to a common ancestor that possessed these proteins. Other molecular data predict that this ancestor was unlikely to perform oxygenic photosynthesis. This evidence, that aerobic respiration has a single origin and may have evolved before oxygen was released to the atmosphere by photosynthetic organisms, is contrary to the textbook viewpoint.

Aerobiosis↗

PubMed1994

Structure and thermal denaturation of crystalline and noncrystalline cytochrome oxidase as studied by infrared spectroscopy.

Fourier-transform infrared spectroscopy has been applied to the study of lipid vesicle-supported two-dimensional crystals and noncrystalline preparations of beef heart cytochrome oxidase. At room temperature, no conformational differences are seen between the noncrystalline and crystalline proteins, whose conformation is shown to consist of ca. 40% alpha-helix, 20% extended structures (including beta-sheet), 17% beta-turns, and 22% open loops plus nonstructured conformations. A novel infrared approach that combines quantitative spectral band decomposition with the study of the thermal behavior of each component has been applied. The procedure allows the independent examination of temperature-induced changes in individual structural elements (alpha-helix, beta-sheet, beta-turns, and unordered). All these reflect, upon heating the protein from 20 to 80 degrees C, a major irreversible thermal event centred at 55-60 degrees C, leading to a molecular state devoid of enzyme activity but with a defined secondary structure; in addition, when the band position, percent area (integrated intensity), and bandwidth of the various amide I components are separately plotted versus temperature, each component is seen to behave in a characteristic way. Thermal denaturation in D2O buffer shows a decrease in nonstructured conformations and an increase in beta-turns without major changes in the proportion of alpha-helix. Temperature-induced changes are not the same in amorphous and crystalline structures, the latter being in general more stable toward the thermal challenge. The above data extend and confirm previous structural studies on cytochrome oxidase using cryo-electron microscopy.

Animals↗

PubMed1994

A second terminal oxidase in Sulfolobus acidocaldarius.

We previously found that the soxABCD operon encodes a quinol oxidase complex in Sulfolobus acidocaldarius and this enzyme was purified and characterized. In this study, we have used a cloning procedure based on the conservation of oxidase sequences and the polymerase chain reaction to isolate a new gene (soxM) encoding a subunit of another terminal oxidase. This terminal oxidase is a fusion between two central components of cytochrome oxidases, subunits I and III. soxM forms a transcriptional unit which is expressed under heterotrophic growth conditions. The corresponding protein was detected by direct protein sequencing in a preparation enriched with a cytochrome absorbing light at 562 nm. This preparation contains a terminal oxidase which is able to oxidize the artificial substrate N,N,N',N'-tetramethyl-p-phenylenediamine. This preparation also contains SoxC, a protein homologous to the mitochondrial cytochrome b, and a Rieske iron-sulphur center. We suggest that SoxM is the core component of a second terminal oxidase complex and that this complex may share a subunit (SoxC) with the SoxABCD complex.

Amino Acid Sequence↗

PubMed1994

Evolution of cytochrome oxidase, an enzyme older than atmospheric oxygen.

Cytochrome oxidase is a key enzyme in aerobic metabolism. All the recorded eubacterial (domain Bacteria) and archaebacterial (Archaea) sequences of subunits 1 and 2 of this protein complex have been used for a comprehensive evolutionary analysis. The phylogenetic trees reveal several processes of gene duplication. Some of these are ancient, having occurred in the common ancestor of Bacteria and Archaea, whereas others have occurred in specific lines of Bacteria. We show that eubacterial quinol oxidase was derived from cytochrome c oxidase in Gram-positive bacteria and that archaebacterial quinol oxidase has an independent origin. A considerable amount of evidence suggests that Proteobacteria (Purple bacteria) acquired quinol oxidase through a lateral gene transfer from Gram-positive bacteria. The prevalent hypothesis that aerobic metabolism arose several times in evolution after oxygenic photosynthesis, is not sustained by two aspects of the molecular data. First, cytochrome oxidase was present in the common ancestor of Archaea and Bacteria whereas oxygenic photosynthesis appeared in Bacteria. Second, an extant cytochrome oxidase in nitrogen-fixing bacteria shows that aerobic metabolism is possible in an environment with a very low level of oxygen, such as the root nodules of leguminous plants. Therefore, we propose that aerobic metabolism in organisms with cytochrome oxidase has a monophyletic and ancient origin, prior to the appearance of eubacterial oxygenic photosynthetic organisms.

Aerobiosis↗

PubMed1994

A role for cholesterol as a structural effector of the nicotinic acetylcholine receptor.

The effects of cholesterol on the protein structure and on the ionic channel activity of purified acetylcholine receptor (AcChR) reconstituted into lipid vesicles have been studied, respectively, by Fourier-transform infrared spectroscopy and by rapid kinetics of cation influx. Reconstitution of the AcChR in asolectin phospholipid vesicles in the absence of either cholesterol or the nonpolar lipids present in crude asolectin extracts results in a considerable loss of the ability of the AcChR to support cation channel function. This functional loss is accompanied by spectral changes in the conformationally-sensitive amide I band of the protein infrared spectrum which are indicative of alteration in the protein secondary structure. Quantitative estimation of such alteration by band-fitting analysis reveals a marked decrease in ordered protein structures such as the alpha-helix and beta-pleated sheet, concomitant with an increase in less ordered structures appearing at 1644 cm-1 in the infrared spectrum. Furthermore, the addition of increasing amounts of cholesterol to the reconstituted bilayer produces a progressive, complete recovery both in the control of cation channel function and in the infrared spectrum. This restoration of AcChR structure and function by cholesterol, however, does not occur when the AcChR is reconstituted in vesicles made from purified egg phosphatidylcholine, thus suggesting that the presence in the reconstituted bilayer of phospholipids other than phosphatidylcholine may be required for cholesterol to exert its modulatory effects.(ABSTRACT TRUNCATED AT 250 WORDS)

Amides↗

PubMed1994

Cytochrome oxidase evolved by tinkering with denitrification enzymes.

The cytochrome bc complex which is encoded by the fixNOPQ operon in Bradyrhizobium japonicum, is the most distant member of the haem-copper cytochrome oxidase family. We have found that its major subunit, FixN, is homologous to the NorB subunit of nitric oxide reductase in a purple bacterium. A second evolutionary link between cytochrome oxidases and denitrification enzymes is the presence of a similar binuclear copper site in cytochrome aa3 (the mitochondrial oxidase) and nitrous oxide reductase. This centre was probably acquired by a primitive FixN-type oxidase, leading to the evolution of the mitochondrial-type oxidase. These links suggest that the oxygen-reducing respiratory chain developed from the anaerobic, denitrifying respiratory system.

Amino Acid Sequence↗