Emile Zuckerkandl

Evolutionary Divergence and Convergence in Proteins

Publisher Summary Informational macromolecules, or semantides, play a unique role in determining the properties of living matter in the perspectives that differ by the magnitude of time required for the processes involved—the short-timed biochemical reaction, the medium-timed ontogenetic event, and the long-timed evolutionary event. Although the slower processes should be broken down into linked faster processes, if one loses sight of the slower processes one also loses the links between the component faster processes. The relative importance of the contributions to evolution of changes in functional properties of polypeptides through their structural modification on the one hand, and of changes in the timing and the rate of synthesis of these polypeptides on the other hand, constitutes a problem that justifies the study of evolution at the level of informational macromolecules. The evaluation of the amount of differences between two organisms as derived from sequences in structural genes or in their polypeptide translation is likely to lead to quantities different from those obtained on the basis of observations made at any other, higher level of biological integration.

Molecules as documents of evolutionary history

Abstract Different types of molecules are discussed in relation to their fitness for providing the basis for a molecular phylogeny. Best fit are the “semantides”, i.e. the different types of macromolecules that carry the genetic information or a very extensive translation thereof. The fact that more than one coding triplet may code for a given amino acid residue in a polypeptide leads to the notion of “isosemantic substitutions” in genic and messenger polynucleotides. Such substitutions lead to differences in nucleotide sequence that are not expressed by differences in amino acid sequence. Some possible consequences of isosemanticism are discussed.

“Homology” in proteins and nucleic acids: A terminology muddle and a way out of it

“Homology” has the precise meaning in biology of “having a common evolutionary origin,” but it also carries the loose meaning of “possessing similarity or being matched.” Its rampant use in the loose sense is clogging the literature on protein and nucleic acid sequence comparisons with muddy writing and, in some cases, muddy thinking In its precise biological meaning, “homology” is a concept of quality. The word asserts a type of relationship between two or more things. Thus, amino acid or nucleotide sequences are either homologous or they are not. They cannot exhibit a particular “level of homology” or “percent homology.” Instead, two sequences possess a certain level of similarity. Similarity is thus a quantitative property. Homologous proteins or nucleic acid segments can range from highly similar to not recognizably similar (where similarity has disappeared through divergent evolution). If using “homology” loosely did not interfere with our thinking about evolutionary relationships, the way in which we use the term would be a rather unimportant semantic issue. The fact is, however, that loose usage in sequence comparison papers often makes it difficult to know the authors intent and can lead to confusion for the reader (and even for the author). There are three common situations in which hazards arise by using “homology” to mean similarity. The first case is the most obvious offense but perhaps the least troublesome. Here an author identifies sequence similarities (calling them homologies) but claims that the sequences being compared are not evolutionarily related. Some awkward moments occur in such a paper, since the author claims both homology (i.e., similarity) and nonhomology (i.e., lack of a common ancestor). Nonetheless, the author’s ideas are likely to be clear since arguments against common ancestry are presented explicitly. A second case is one in which an author points out similarities (again called homologies) but does not address the issue of evolutionary origins. The reader, seeing the term “homology,” may infer that the author is postulating coancestry when that is not the authors intent. The final case occurs most frequently and is the most subtle and therefore most troublesome. Here, similarities (called homologies) are used to support a hypothesis of evolutionary homology. In this case, the two meanings of homology seem to overlap, and it is almost inevitable that the thinking of author and reader alike will be intrusively distorted as follows. Similarity is relatively straightforward to document. In comparing sequences, a similarity can take the form of a numerical score (O/o amino acid or nucleotide positional identity, in the simplest approach) or of a probability associated with such a score. In comparisons of three-dimensional structures, a typical numerical description is root-mean-square positional deviation between compared atomic positions. A similarity, then, can become a fully documented, simple fact. On the other hand, a common evolutionary origin must usually remain a hypothesis, supported by a set of arguments that might include sequence or three-dimensional similarity. Not all similarity connotes homology but that can be easily overlooked if similarities are called homologies. Thus, in this third case, we can deceive ourselves into thinking we have proved something substantial (evolutionary homology) when, in actuality, we have merely established a simple fact (a similarity, mislabeled as homology). Homology among similar structures is a hypothesis that may be correct or mistaken, but a similarity itself is a fact, however it is interpreted. We believe that the concepts of evolutionary homology and sequence or three-dimensional similarity can be kept distinct only if they are referred to with different words. We therefore offer the following recommendations: *Sequence similarities (or other types of similarity) should simply be called similarities. They should be documented by appropriate statistical analysis. In writing about sequence similarities the following sorts of terms might be used: a level or degree of similarity; an alignment with optimized similarity; the % positional identity in an alignment; the probability associated with an alignment. *Homology should mean “possessing a common evolutionary origin” and in the vast majority of reports should have no other meaning. Evidence for evolutionary homology should be explicitly laid out, making it clear that the proposed relationship is based on the level of observed similarity, the statistical significance of the similarity, and possibly other lines of reasoning. One could argue that the meaning of the term “homology” is itself evolving. But if that evolution is toward vagueness and if it results in making our scientific discourse unclear, surely we should intervene. With a collective decision to mend our ways, proper usage would soon become fashionable and therefore easy. We believe that we and our scientific heirs would benefit significantly.

Standardized nomenclature for Alu repeats

1 Human Genome Center, L-452, Biology and Biotechnology Research Program, Lawrence Livermore National Laboratory, P.O. Box 808, Livermore, CA 94551, USA 2 Department of Biochemistry and Molecular Biology, Center for Human and Molecular Genetics, Louisiana State University Medical Center, 1901 Perdido St., New Orleans, LA 70112, USA 3 Department of Chemistry, University of California at Davis, Davis, CA 95616, USA 4 Linus Pauling Institute of Science and Medicine, 440 Page Mill Road, Palo Alto, CA 94306, USA 5 Centre de Recherche, Hopital Ste-Justine, Departement de Pediatrie, Universite de Montreal, Montreal, Quebec, Canada H3T 1C5 6 Section of Molecular and Cell Biology, University of California at Davis, Davis, CA 95616, USA 7 Institute of Molecular Medical Sciences, 460 Page Mill Road, Palo Alto, CA 94306, USA

Evolutionary processes and evolutionary noise at the molecular level. I. Functional density in proteins.

SummaryThe distinction between molecular sites that mainly carry out general functions and sites committed to specific functions is analyzed, notably in terms of different evolutionary variabilities. Functional density is defined as the proportion of sites involved in specific functions. Weighted functional density, by representing the relative variability at specific-function sites is to some extent a measure of the specificity of molecular interactions. The relationship between general- and specific-function sites on the one hand and the covarions of Fitch on the other is discussed. The functional −degeneracy− of amino acids is described as increasing the interdependence of general functions. It is predicted that proteins that do not possess general-function sites besides their specific-function sites tend to −freeze− their primary structure, according to an evolutionary process that is an autocatalytic function of the decrease in site variability. This limits the use of weighted functional density as an indicator of the overall degree of interaction specificity of a protein to values that are not close to unity.

The appearance of new structures and functions in proteins during evolution.

SummaryThe likelihood of a de novo generation of classes of efficient proteins through neoformation of DNA, through modification of expressed DNA, and through modification of nonexpressed DNA is examined. So is the likelihood that newly formed inefficient enzymes be turned into efficient enzymes. The conclusions are that neither gene duplicates nor dormant genes represent promising materials for a de novo generation of protein classes, that (with exceptions) such generation is unlikely to have taken place in recent evolution, that new structural genes must nearly consistently derive from preexisting structural genes, and that new functions can be evolved only on the basis of old proteins. Conditions of protein evolution in prokaryotes suggest that the saltatory formation of protein classes is as unlikely in prokaryotes as in eukaryotes. Data on the history of a few protein classes are reviewed to illustrate the preceding inferences. The analysis leads to the hypothesis that most protein classes originated before the major elements of the translation apparatus of modern cells were fully evolved. If simple sequence DNA is turned into structural genes by evolution, this process (again with exceptions) is considered to have taken place only at that very remote period. A polyphyletic origin of proteins is thought to date back to the same era. It is proposed that the development of genic multiplicity and of marked structural and functional diversity of proteins may have come about in the earliest cells primarily through the independent generation of structurally different polymerases in different protocells, followed by cell conjugation and the subsequent use by enriched cells of supernumerary types of polymerase for evolving further functions. Functional growth, as it took place at early times, is briefly discussed as well as functional change. The foundations for new functional developments in old proteins are analyzed. In considering the evolutionary recovery of lost functions, aspects of cell differentiation and gene regulation are linked with the evolutionary picture. The distinction between eurygenic and stenogenic control of gene activity is used. Next to gene deletion, cell and tissue deletion is held to be an event of general evolutionary significance, through cell and tissue origination that presumably accompanies the restoration of a lost molecular function.

On the Molecular Evolutionary Clock

SummaryThe conceptual framework surrounding the origin of the molecular evolutionary clock and circumstances of this origin are described. In regard to the quest for the best available molecular clocks, a return to protein clocks is conditionally recommended. On the basis of recent data and certain considerations, it is pointed out that the realm of neutrality in evolution is probably less extensive than is now commonly thought, in the three distinct senses of the term neutrality—neutrality as nonfunctionality of mutations, neutrality as equifunctionality of mutations, and neutrality as a mode of fixation of mutations. The possibility is raised that complex sets of interacting components forming a system that is bounded with respect to its environment may quite generally display an intrinsic trend to a quasi-clockwise evolutionary behavior.

Controller-gene diseases: The operon model as applied to β-thalassemia, familial fetal hemoglobinemia and the normal switch from the production of fetal hemoglobin to that of adult hemoglobin

It is shown that the concepts evolved by Jacob & Monod (1961a,b) as an outcome of their studies on the lactose system in Escherichia coli can be applied to a mammalian system, namely to the synthesis of human hemoglobin polypeptide chains, in a way that appears consistent with the known facts. In the system involving the β, δ- and γ-chain genes the intervention of two distinct operators is postulated. Reasons are given why the regulator gene involved in the control of γ-chain production is probably not identical with any of the known structural hemoglobin chain genes. The activity of the γ-chain gene is considered to be submitted to a double control, namely on the one hand to an interplay of regulator genes and operators, and on the other hand to an inducer that reacts with the product of a regulator gene. Physiological conditions are thought to influence the state of activity of the γ-chain gene via the inducer. Familial fetal hemoglobinemia is considered as an operator-negative mutation. Other mutations are examined that would lead to a phenotypically comparable expression. The occurrence of some as yet unobserved mutations, such as familial adult hemoglobinemia in the fetus, is predicted. In opposition to suggestions that have been made, β-thalassemia, in typical cases, appears to be attributable neither to a mutation in the affected structural gene, nor to a mutation in an operator-like factor. It might be attributable to a translocation (or inversion), or to a “silent mutation” in an unknown structural gene located between the β-chain gene and its operator. The probable role of controller-gene diseases in evolution is pointed out.

Mutational trends and random processes in the evolution of informational macromolecules

Abstract On the basis of molecular phylogenetic trees, the frequency of base substitutions in the gene and of amino-acid substitutions in the corresponding protein are studied for globin and cytochrome c . An analysis of directional trends in base substitutions separately at the first and at the second coding positions shows that they are unlikely to express intrinsic mutational trends. On the other hand the data are in part compatible with the expression in evolution of random mutational events. On the level of amino-acid substitutions it is noted that, on the average, the more frequent amino-acid residues tend to get rarer and the rarer ones more frequent. Thereby a random process is again suggested, though not established, in molecular evolution.

Why so many noncoding nucleotides? The eukaryote genome as an epigenetic machine.

It is recalled that dispensability of sequences and neutral substitution rate must not be construed to be markers of nonfunctionality. Different aspects of functionality relate to differently-sized nucleotide communities. At the time cells became nucleated, a boom of epigenetic processes led to uses of DNA that required many more nucleotides operating collectively than do functions definable in terms of classical genetics. Each order of magnitude of nucleotide plurality was colonized by functions germane to that order. The eukaryote genome became a great epigenetic machine. Sequences of different levels of nucleotide plurality are briefly discussed from the point of view of their functional relevance. By their activities as both transcribed genes and cis-acting repeats, SINEs and LINEs are the principal link between genetic and epigenetic processes. SINEs can act as local repeats to produce position effect variegation (PEV) in a nearby gene. PEV may thus represent a general method of overall transcriptional regulation at the level of cell collectivities. When tracking the scale dependence of nucleotide function, one finds the 100 kb order of nucleotide plurality to provide epigenetically the basis at once for PEV, imprinting, and cell determination, with sectorial repressibility a trait common to the three. In sectorial repressibility, introns may play a structural role favoring the stability of higher-order chromatin structures. At that level of nucleotide involvement, nonconserved nonhomologous nonprotein-coding sequences may often play the same structural roles. In addition, genomic distance per se – and, therefore, the mass of intervening nucleotides – can have functional effects. Distances between enhancers and promoters need to be probed in this respect. At the 1000 kb level of nucleotide function, attention is focused on the formation of centromeres. It is one of the levels of nucleotide plurality per function where specificity in the generation of DNA/protein complexes seems to depend more upon the structural fit among factors than upon the DNA sequence. This circumstance may explain in part the prevailing difficulty in recognizing the functional nature of sequences among non-protein-coding nucleotide arrays and the propensity among investigators to tag the majority of DNA sequences in higher organisms as functionally meaningless. Noncoding DNA often may not be ‘selected’ as an appropriate niche for a certain function, but be ‘elected’ in that capacity by a group of factors, as a preexisting sequence that is only now called upon to serve. Much of the non-protein-coding DNA may thus be only conditionally functional and in fact may never be elected to functions at a high level of nucleotide plurality. Eukaryotes are composites, at different levels of this plurality, of the functional and the nonfunctional, as well as of the conditionally functional and the outright functional. Thus, a sequence that is nonfunctional at one level of nucleotide plurality may participate in a functional sequence at a more inclusive level. In the end, every nucleotide is at least infinitesimally functional if, for metabolic and developmental reasons, the chromatin mass as such becomes a selectable entity. Given the scale dependence of nucleotide function, large amounts of ‘junk DNA’, contrary to common belief, must be assumed to contribute to the complexity of gene interaction systems and of organisms.