Similar amino acid sequences: chance or common ancestry?

The systemic comparison of every newly determined amino acid sequence with all other known sequences may allow a complete reconstruction of the evolutionary events leading to contemporary proteins. But sometimes the surviving similarities are so vague that even computer-based sequence comparisons procedures are unable to validate relationships. In other cases similar sequences may appear in totally alien proteins as a result of mere chance or, occasionally, by the convergent evolution of sequences with special properties.

Angiotensinogen is related to the antitrypsin-antithrombin-ovalbumin family.

The recently reported amino acid sequence of rat angiotensinogen was subjected to a computer-assisted search for homology with known sequences stored in a data bank and found to be significantly related to that of plasma alpha 1-antitrypsin, itself a member of a family that includes antithrombin III and ovalbumin. An alignment of the four sequences shows indisputably the common ancestry of all four proteins.

Platelet and plasma fibrinogens are identical gene products.

Human platelet fibrinogen has been shown to be indistinguishable from plasma fibrinogen by a variety of criteria, including subunit composition and cross-linking characteristics as judged by sodium dodecyl sulfate-gel electro phoresis, quantitative amino terminal analysis of fibrin, and the amino acid compositions of fibrinopeptides released from platelet and plasma fibrinogens is products of the same gene or genes.

Evolution of immunoglobulin polypeptide chains: carboxy-terminal of an IgM heavy chain.

The dipeptide sequence at the carboxy-terminal of a heavy (micro) chain from a human macroglobulin ( IgM) is tyrosylcysteine, although the reverse sequence, cysteinyltyrosine, has not been rigorously excluded. The presence of cysteine at the carboxy-terminal was predicted from a recognition of the chemical homologies among the polypeptide chains of immunoglobulins, and their probable evolutionary origin.

Antibody active sites and immunoglobulin molecules.

In order to obtain detailed information about the relationship between structure and function in antibody molecules, a method called affinity labeling has been devised to attach chemical labels specifically to amino acid residues in the active sites of antibody molecules. With antibodies to three different haptens, highly specific labeling of the active sites has been achieved. Tyrosine residues on both heavy and light polypeptide chains have been labeled in a molar ratio close to 2:1, and labels on the two chains are equally specific to the active sites. Peptide fragmentation studies of the labeled chains of one antibody system have shown that: (i) within 25 amino acid residues of the labeled tyrosine on either chain, substantial chemical heterogeneity exists among different antibody molecules of the same specificity; and (ii) the labeled peptide fragments from both chains are very similar in physicochemical characteristics, including average size, heterogeneity, and unusual hydrophobicity. These experimental results have led us to the view that a particular region of the heavy chain and a particular region of the light chain are utilized to construct the active sites of the three different antibodies, differences in specificity arising from chemical perturbations in these two regions. Correlated structural studies of affinity-labeled antibodies and of the homogeneous light chains (Bence Jones proteins) and heavy chains produced in multiple myeloma may permit the identification of these special active-site regions. The view that active sites of different specificity are chemical perturbations of a particular region of the antibody molecule has a possible close analogue in enzyme systems, particularly among the esterases. The marked chemical similarities we have observed between the active site regions of heavy and light chains indicate to us that chemical homologies, but not identities, exist between the chains. This is reinforced by recently obtained amino acid sequence data which reveal homologies between the two chains near their carboxyl-terminals. These results indicate that the structural genes which code for the synthesis of heavy and light chains are related, presumably having arisen from some common ancestral gene during evolution. This conclusion strongly suggests that both heavy and light chains determine antibody specificity, and has important implications for the still-unknow mechanisms of antibody biosynthesis.

Light chains of rabbit immunoglobulin: assignment to the kappa class.

Normal rabbit gamma globulin was reduced under conditions presumed to break only interchain disulfide bridges, and the reduiced product was then blocked with C(14)-iodoacetamide. The light chains were separated from the heavy chains and subjected to peptic digestion. Two radioactive peptides were recovered from the digest. The peptides are apparently overlapping and represent the carboxyterminuis. Comparison of this region in the rabbit light chains with the corresponding amino acid sequences in various mouse and human light chains indicates that the rabbit light chains are of the (K)-class.

Influence of calcium ion on the binding of fibrin amino terminal peptides to fibrinogen.

The affinity of the amino terminal tetrapeptide of the beta chain of fibrin, Gly-His-Arg-Pro, for fibrinogen dramatically increases in the presence of 2 millimolar calcium ion. In contrast, there is no significant increase in the affinity of peptides beginning with the amino terminal sequence of the fibrin alpha chain, Gly-Pro-Arg, in the presence of calcium ions, although the number of binding sites increases. In the latter case, the increased number of sites is due to the alpha chain analogs binding to the site ordinarily occupied by the beta chain analogs. These results indicate that structures at the amino terminus of the fibrin beta chain play a more important role in fibrin polymerization when calcium ions are present.

Gibbon fibrinopeptides: identification of a glycine-serine allelism at position B-3.

The fibrinopeptides A and B of the gibbon (an Asian ape) have been characterized and their relation to other primate types examined. An allelic situation was discovered at location B-3; two of the gibbons studied had both glycine and serine at that position, whereas four others were homozygous for glycine.

Determining divergence times of the major kingdoms of living organisms with a protein clock.

Amino acid sequence data from 57 different enzymes were used to determine the divergence times of the major biological groupings. Deuterostomes and protostomes split about 670 million years ago and plants, animals, and fungi last shared a common ancestor about a billion years ago. With regard to these protein sequences, plants are slightly more similar to animals than are the fungi. In contrast, phylogenetic analysis of the same sequences indicates that fungi and animals shared a common ancestor more recently than either did with plants, the greater difference resulting from the fungal lineage changing faster than the animal and plant lines over the last 965 million years. The major protist lineages have been changing at a somewhat faster rate than other eukaryotes and split off about 1230 million years ago. If the rate of change has been approximately constant, then prokaryotes and eukaryotes last shared a common ancestor about 2 billion years ago, archaebacterial sequences being measurably more similar to eukaryotic ones than are eubacterial ones.

URF6, last unidentified reading frame of human mtDNA, codes for an NADH dehydrogenase subunit.

The polypeptide encoded in URF6, the last unassigned reading frame of human mitochondrial DNA, has been identified with antibodies to peptides predicted from the DNA sequence. Antibodies prepared against highly purified respiratory chain NADH dehydrogenase from beef heart or against the cytoplasmically synthesized 49-kilodalton iron-sulfur subunit isolated from this enzyme complex, when added to a deoxycholate or a Triton X-100 mitochondrial lysate of HeLa cells, specifically precipitated the URF6 product together with the six other URF products previously identified as subunits of NADH dehydrogenase. These results strongly point to the URF6 product as being another subunit of this enzyme complex. Thus, almost 60% of the protein coding capacity of mammalian mitochondrial DNA is utilized for the assembly of the first enzyme complex of the respiratory chain. The absence of such information in yeast mitochondrial DNA dramatizes the variability in gene content of different mitochondrial genomes.

Simian sarcoma virus onc gene, v-sis, is derived from the gene (or genes) encoding a platelet-derived growth factor.

The transforming protein of a primate sarcoma virus and a platelet-derived growth factor are derived from the same or closely related cellular genes. This conclusion is based on the demonstration of extensive sequence similarity between the transforming protein derived from the simian sarcoma virus onc gene, v-sis, and a human platelet-derived growth factor. The mechanism by which v-sis transforms cells could involve the constitutive expression of a protein with functions similar or identical to those of a factor active transiently during normal cell growth.

Structure and expression of a complementary DNA for the nuclear coded precursor of human mitochondrial ornithine transcarbamylase.

Most mitochondrial proteins are encoded in the nucleus and are translated on free cytoplasmic ribosomes as larger precursors containing amino-terminal "leader" sequences, which are removed after the precursors are taken up by mitochondria. We have deduced the complete primary structure of the precursor of a human mitochondrial matrix enzyme, ornithine transcarbamylase (OTC), from the nucleotide sequence of cloned complementary DNA. The amino-terminal leader peptide of OTC is 32 amino acids in length and contains four arginines but no acidic residues. Cleavage of the leader peptide from the "mature" protein occurs between glutamine and asparagine residues. The sequence of mature human OTC resembles that of the subunits of both OTC and aspartate transcarbamylase from Escherichia coli. The biological activity of the cloned OTC complementary DNA was tested by joining it with SV40 (an animal virus) regulatory elements and transfecting cultured HeLa cells, which do not normally express OTC. Both the precursor and mature forms of the OTC subunit were identified; in stable transformants, enzymatic activity was also detected.

Convergent evolution: the need to be explicit.

Convergence as a phenomenon in molecular evolution is an issue that confuses many discussions. Often the problem is that not enough care is taken to state exactly what kind of convergence one has in mind. Functional and mechanistic convergence are both common, and some structural convergence has probably occurred, but a convincing case for genuine sequence convergence has yet to be made.

Similar amino acid sequences revisited.

The rapid accumulation of protein sequences, many bearing unexpected resemblances to each other, is providing a new perspective on evolution.

Evolution by acquisition: the case for horizontal gene transfers.

One of the most debated questions in the field of molecular evolution is the possible role of horizontal transfer in evolution. Of all the claims that have been made over the years, those reporting transfers between eukaryotes and prokaryotes are the most controversial. Here we present the cases for and against several such possible gene acquisitions.

Evolutionary anomalies among the aminoacyl-tRNA synthetases.

Unexpected relationships among the various aminoacyl-tRNA synthetases continue to be uncovered. The question arises - is this mainly the result of promiscuous exchange, or is the confusion really a reflection of the differential loss of past duplications? Phylogenetic analysis may yet provide the answer.

Three-dimensional structural studies on fragments of fibrinogen and fibrin.

Fibrinogen is a 340 kDa glycoprotein found in the blood plasma of all vertebrates. It is transformed into a fibrin clot by the action of thrombin. Recent X-ray structures of core fragments of both fibrinogen and fibrin have revealed many details about this polymerization event. These include structures of a 30 kDa recombinant gammaC domain, an 86 kDa fragment D from human fibrinogen and a cross-linked double-D fragment from fibrin.

Amino acid sequence of the carboxy-terminal cyanogen bromide peptide of the human fibrinogen beta-chain: homology with the corresponding gamma-chain peptide and presence in fragment D.

The carboxy-terminal cyanogen bromide fragment of the human fibrinogen beta-chain has been isolated and its structure determined. It is a nonapeptide with the sequence Lys-Ile-Arg-Pro-Phe-Phe-Pro-Gln-Gln and is homologous with a portion of the carboxy-terminal cyanogen bromide fragment of the gamma-chain. The peptide has also been isolated in full yield from cyanogen bromide digests of the plasmin-derived fragment D, indicating that the carboxy-terminal region of the beta-chain is resistant to plasmin digestion. In contrast, a small portion of the corresponding gamma-chain carboxy-terminal region was missing in the same fragment D.

Amino acid sequences of lamprey fibrinopeptides A and B and characterizations of the junctions split by lamprey and mammalian thrombins.

The amino acid sequences of the fibrinopeptides A and B from lamprey fibrinogen have been determined. The fibrinopeptide A is the shortest fibrinopeptide ever isolated, being comprised of only six amino acids. The fibrinopeptide B, on the other hand, is the largest fibrinopeptide characterized to date, having 36 amino acid residues and a cluster of covalently bound carbohydrate. As reported previously, lamprey fibrinogen is readily clotted by mammalian thrombins, but only the fibrinopeptide B is released during the process. Lamprey fibrinopeptide A is not released by mammalian thrombins and could only be removed with the use of lamprey thrombin. Firm proof that the lamprey fibrinopeptides A and B are the amino segments of the alpha and beta-chains respectively was obtained by a series of stepwise degradations on lamprey fibrinogen and lamprey fibrins produced in turn by the action of mammalian thrombin (fibrin B) and lamprey thrombin (fibrin A). These studies were supplemented by stepwise degradations on the individual Aalpha and Bbeta-chains. It the case of the lamprey Aalpha-chain it was also possible to release the 6-residue fibrinopeptide A from the isolated chain with lamprey thrombin and demonstrate that the newly exposed amino-terminal sequence begins with the Gly-Pro-Arg sequence characteristic of mammalian fibrin alpha-chains. In fact, the sequences on the fibrin side of both of the junctions split by thrombin(s) are highly conserved and virtually identical with those found in mammalian alpha and beta-chains.