Characterization of the N-linked high-mannose oligosaccharides of the insulin pro-receptor and mature insulin receptor subunits.

An interesting pattern in the genetic code was reported previously [Blalock & Smith (1984) Biochem. Biophys. Res. Commun. 121, 203-207]. In the 5'-to-3' direction, codons for hydrophilic and hydrophobic amino acids are generally complemented by codons for hydrophobic and hydrophilic amino acids respectively. The average tendency of codons for 'unchanged' (slightly hydrophilic) amino acids was to be complemented by codons for 'unchanged' amino acids. We now show that the same pattern results when the complementary codon is read in the 3'-to-5' direction. This pattern is further shown to result in the interaction of peptides specified by complementary RNAs regardless of whether the amino acids are assigned in the 5'-to-3' or the 3'-to-5' direction. Here we demonstrate that peptides specified by complementary RNAs bind to each other with specificity and high affinity.     

Complementary hydropathy and the evolution of interacting polypeptides

Summary It has been shown that codons coding for strongly hydrophilic amino acids are complemented by codons that code for strongly hydrophobic ones, leading to a hypothesis stating that peptides thus encoded should interact. Though the principle has been validated in a number of experimental models, its general applicability has been questioned. I have discussed this principle, showing that the correlation between coding and noncoding strand amino acids was maintained, indeed slightly improved, when weighted averages based on codon usage tables were used to determine noncoding strand amino acid hydropathies. The coding capacity of the noncoding strand and its content of open reading frames were also discussed. Another point of contention that was afforded further clarification is the chemical plausibility of interactions between hydrophobic and hydrophilic amino acids implicit in this concept. The extension of complementary domains was also dealt with. Finally, I have discussed what I called the evolutionary drift of primary structure, and I showed as an example that though nucleotide sequences coding for the substance K receptor bear little resemblance to the inverse complement of that which codes for the SK peptide, a peptide spanning residues 130–139 is hydropathically very similar to that predicted from such an inverse complement.     

Role of mutational bias and natural selection on genome-wide nucleotide bias in prokaryotic organisms

Correlations between genomic GC contents and amino acid frequencies were studied in the homologous sequences of 12 eubacterial genomes. Results show that amino acids encoded by GC-rich codons increases significantly with genomic GC contents, whereas opposite trend was observed in case of amino acids encoded by GC-poor codons. Further studies show all the amino acids do not change in the predicted direction according to their genomic GC pressure, suggesting that protein evolution is not entirely dictated by their nucleotide frequencies. Amino acid substitution matrix calculated among hydrophobic, amphipathic and hydrophilic amino acid groups' shows that amphipathic and hydrophilic amino acids are more frequently substituted by hydrophobic amino acids than from hydrophobic to hydrophilic or amphipathic amino acids. This indicates that nucleotide bias induces a directional changes in proteome composition in such a way that underwent strong changes in hydropathy values. In fact, significant increases in hydrophobicity values have also been observed with the increase of genomic GC contents. Correlations between GC contents and amino acid compositions in three different predicted protein secondary structures show that hydropathy values increases significantly with GC contents in aperiodic and helix structures whereas strand structure remains insensitive with the genomic GC levels. The relative importance of mutation and selection on the evolution of proteins have been discussed on the basis of these results.     

Intrinsic disorder in protein sense‐antisense recognition

In addition to the well‐established sense‐antisense complementarity abundantly present in the nucleic acid world and serving as a basic principle of the specific double‐helical structure of DNA, production of mRNA, and genetic code‐based biosynthesis of proteins, sense‐antisense complementarity is also present in proteins, where sense and antisense peptides were shown to interact with each other with increased probability. In nucleic acids, sense‐antisense complementarity is achieved via the Watson‐Crick complementarity of the base pairs or nucleotide pairing. In proteins, the complementarity between sense and antisense peptides depends on a specific hydropathic pattern, where codons for hydrophilic and hydrophobic amino acids in a sense peptide are complemented by the codons for hydrophobic and hydrophilic amino acids in its antisense counterpart. We are showing here that in addition to this pattern of the complementary hydrophobicity, sense and antisense peptides are characterized by the complementary order‐disorder patterns and show complementarity in sequence distribution of their disorder‐based interaction sites. We also discuss how this order‐disorder complementarity can be related to protein evolution.     

Distribution of mapping points of 20 amino acids in the tetrahedral space

Summary Based on the genetic codes and a simple theorem for the geometrical property of the regular tetrahedron, each amino acid is mapped onto a unique point in a 3-dimensional tetrahedral space. The distribution of the 20 mapping points for 20 amino acids is studied in detail. It is found that the mapping points for the hydrophobic and hydrophilic amino acids are distributed at distinct regions in the 3-dimensional space. A plane separating the two kinds of points satisfactorily based on the Fisher's algorithm has been calculated. It is shown that the codons coding for the hydrophobic amino acids are constituted dominantly by the bases of keto group, i.e., G and T. While the codons coding for the hydrophilic amino acids are constituted dominantly by the bases of amino group, i.e., A and C. The biological implication of the mapping points and the separating plane has been discussed in some details.     

Biological implications of complementary hydropathy of amino acids

The principle of complementary hydropathy predicts that peptides coded for by opposing DNA strands will bind one another because highly hydrophilic amino acids will be complemented by hydrophobic ones and vice versa. This paper provides the chemical plausibility for such interactions. It is suggested that exons coding for interacting peptides were juxtaposed and co-evolved together. Present day genes are no longer thus arranged because of duplications and exon shuffling.     

Differential effect of amino acid residues on the stability of double helices formed from polyribonucleotides and its possible relation to the evolution of the genetic code

Summary The interaction of amino acid residues with polyribonucleotides was characterized by measurements of melting temperatures (t_m) for poly(A) poly(U) and poly(I)poly(C) as functions of the concentrations of various amino acid amides. The amides of hydrophilic amino acids lead to a continuous increase of tm with increasing concentration, whereas amides of hydrophobic amino acids induce a decrease of t_m at low concentrations (1 mM) followed by an increase at higher concentrations. Analysis of the data by a simple site model provides the affinity of each ligand for the double helix relative to that for the single strands. This parameter decreases in the order Ala>Gly>Ser>Asn>Pro>Met, Val>Ile, Leu for poly(A) poly(U) and Ala, Gly, Ser>Asn>Pro>Val>Ile, Met, Leu for poly(I)poly(C). The special effects of hydrophobic amino acids may be related to the similarity of the codons for these amino acids. A simple model for assignment of codons to amino acids is proposed.     

Genetic Code Evolution Started with the Incorporation of Glycine,Followed by Other Small Hydrophilic Amino Acids

We propose that glycine was the first amino acid to be incorporated into the genetic code, followed by serine, aspartic and/or glutamic acid—small hydrophilic amino acids that all have codons in the bottom right-hand corner of the standard genetic code table. Because primordial ribosomal synthesis is presumed to have been rudimentary, this stage would have been characterized by the synthesis of short, water-soluble peptides, the first of which would have comprised polyglycine. Evolution of the code is proposed to have occurred by the duplication and mutation of tRNA sequences, which produced a radiation of codon assignment outwards from the bottom right-hand corner. As a result of this expansion, we propose a trend from small hydrophilic to hydrophobic amino acids, with selection for longer polypeptides requiring a hydrophobic core for folding and stability driving the incorporation of hydrophobic amino acids into the code.     

A review of adsorption of amino acids on minerals: Was it important for origin of life?

Minerals more readily adsorb amino acids with charged R groups than uncharged R groups, so that the incorporation of amino acids with charged R groups into peptides would be more frequent than for amino acids with uncharged R groups. However, 74% of the amino acids in the proteins of modern organisms contain uncharged R groups. Thus, what could have been the mechanisms that produced peptides/proteins with more amino acids with uncharged R groups than precursors with charged R groups? Should we expect the composition of amino acids adsorbed on minerals to be similar to those of present proteins? Was the adsorption of amino acids on minerals important for the origin of life? The lipid world offers an alternative view of origin of life. Liposomes contributed to elongation of peptides as well as select hydrophobic amino acids and peptides. These experiments could be showing the mechanism, which hydrophobic amino acids have been selected. However, liposomes have no influence on the stereoselectivity in the oligomerization of amino acids. In the present paper, several other mechanisms are also discussed that could produce peptides with a greater proportion of amino acids with uncharged R groups.     

A general model of the P2 protein of peripheral nervous system myelin based on secondary structure predictions, tertiary folding principles, and experimental observations

The amino acid sequence of the P2 protein of peripheral myelin was analyzed with regard to regions of probable alpha-helix, beta-structure, beta-turn, and unordered conformation by means of several algorithms commonly used to predict secondary structure in proteins. Because of the high beta-sheet content and virtual absence of alpha-helix shown by the circular dichroic spectra of the protein, a bias was introduced into the algorithms to favor the beta-structure over the alpha-helical conformation. In order to define those beta-sheet residues that could lie on the external hydrophilic surface of the protein and those that could lie in its hydrophobic interior, the predicted beta-strands were examined for charged and uncharged amino acids located at alternating positions in the sequence. The sequential beta-strands in the predicted secondary structure were then ordered into beta-sheets and aligned according to generally accepted tertiary folding principles and certain chemical properties peculiar to the P2 protein. The general model of the P2 protein that emerged was a "Greek key" beta-barrel, consisting of eight antiparallel beta-strands with a two-stranded ribbon of antiparallel beta-structure emerging from one end. The model has an uncharged, hydrophobic core and a highly hydrophilic surface. The two Cys residues, which form a disulfide, occur in a loop connecting two adjacent antiparallel strands. Two hydrophilic loops, each containing a cluster of acidic residues and a single Phe, protrude from one end of the molecule. The general model is consistent with many of the properties of the actual protein, including the relatively weak nature of its association with myelin lipids and the positions of amino acid substitutions. Alternative beta-strand orderings yield three specific models having different interstrand connections across the barrel ends.     

Second codon positions of genes and the secondary structures of proteins. Relationships and implications for the origin of the genetic code

The nucleotide frequencies in the second codon positions of genes are remarkably different for the coding regions that correspond to different secondary structures in the encoded proteins, namely, helix, beta-strand and aperiodic structures. Indeed, hydrophobic and hydrophilic amino acids are encoded by codons having U or A, respectively, in their second position. Moreover, the beta-strand structure is strongly hydrophobic, while aperiodic structures contain more hydrophilic amino acids. The relationship between nucleotide frequencies and protein secondary structures is associated not only with the physico-chemical properties of these structures but also with the organisation of the genetic code. In fact, this organisation seems to have evolved so as to preserve the secondary structures of proteins by preventing deleterious amino acid substitutions that could modify the physico-chemical properties required for an optimal structure.     

Differential effect of amino acid residues on the stability of double helices formed from polyribonucleotides and its possible relation to the evolution of the genetic code

The interaction of amino acid residues with polyribonucleotides was characterized by measurements of melting temperatures (tm) for poly(A).poly(U) and poly(I).poly(C) as functions of the concentrations of various amino acid amides. The amides of hydrophilic amino acids lead to a continuous increase of tm with increasing concentration, whereas amides of hydrophobic amino acids induce a decrease of tm at low concentrations (approximately 1 mM) followed by an increase at higher concentrations. Analysis of the data by a simple site model provides the affinity of each ligand for the double helix relative to that for the single strands. This parameter decreases in the order Ala greater than Gly greater than Ser greater than Asn greater than Pro greater than Met, Val greater than Ile, Leu for poly(A).poly(U) and Ala, Gly, Ser greater than Asn greater than Pro greater than Val greater than Ile, Met, Leu for poly(I).poly(C). The special effects of hydrophobic amino acids may be related to the similarity of the codons for these amino acids. A simple model for assignment of codons to amino acids is proposed.     

三联体密码子在mRNA二级结构中的分布

对编码成熟肽的ｍＲＮＡ二级结构的分析显示,每个密码子在ｍＲＮＡ二级结构中的位置有一定的倾向性,这种倾向性似乎与相应氨基酸的构象性质相一致。大多数编码疏水氨基酸的密码子位于ｍＲＮＡ二级结构中较稳定的茎区;反之,大多数编码亲水氨基酸的密码子位于柔性的环区。这个结果支持了最近得到的关于ｍＲＮＡ与蛋白质之间存在丰三维结构信息传递的结论。     

Correlation between amino acid residues converted by RNA editing and functional residues in protein three-dimensional structures in plant organelles

Background

In plant organelles, specific messenger RNAs (mRNAs) are subjected to conversion editing, a process that often converts the first or second nucleotide of a codon and hence the encoded amino acid. No systematic patterns in converted sites were found on mRNAs, and the converted sites rarely encoded residues located at the active sites of proteins. The role and origin of RNA editing in plant organelles remain to be elucidated.

Results

Here we study the relationship between amino acid residues encoded by edited codons and the structural characteristics of these residues within proteins, e.g., in protein-protein interfaces, elements of secondary structure, or protein structural cores. We find that the residues encoded by edited codons are significantly biased toward involvement in helices and protein structural cores. RNA editing can convert codons for hydrophilic to hydrophobic amino acids. Hence, only the edited form of an mRNA can be translated into a polypeptide with helix-preferring and core-forming residues at the appropriate positions, which is often required for a protein to form a functional three-dimensional (3D) structure.

Conclusion

We have performed a novel analysis of the location of residues affected by RNA editing in proteins in plant organelles. This study documents that RNA editing sites are often found in positions important for 3D structure formation. Without RNA editing, protein folding will not occur properly, thus affecting gene expression. We suggest that RNA editing may have conferring evolutionary advantage by acting as a mechanism to reduce susceptibility to DNA damage by allowing the increase in GC content in DNA while maintaining RNA codons essential to encode residues required for protein folding and activity.     

Construction of genetic code from evolutionary stability

The construction of the genetic code is investigated based on a stability principle. The concept and formulation of mutational deterioration (MD) of the genetic code is proposed. It is proved that the degeneracies of codon multiplets obey the rule to best resist MD. The MD for each ideal multiplet of codons is expressed by four parameters and it takes on a minimum value for real distributions of codons in the multiplet. Then the global mutational deterioration (GMD) of code table is calculated and the minimal code is deduced. The domain-like distribution of hydrophobic and hydrophilic amino acids on the genetic code is explained from the minimization of GMD. It is demonstrated that the standard code is approximately GMD-minimal. By introducing some constraints that are related to the initial condition of the system, we have deduced the standard genetic code from the minimization of GMD. The minimization shows the general trend of evolutionary process to some stable state while the constraints reflect a 'frozen accident.' Many deviant codon assignments are also explained through MD minimization assuming the changeable degrees of degeneracies for some multiplets. So, a possible answer to the question of "Why are synonymous codons and amino acids distributed in the code table just as they are?" is given.     

Evolution of the genetic code.

Comparative path lengths in amino acid biosynthesis and other molecular indicators of the timing of codon assignment were examined to reconstruct the main stages of code evolution. The codon tree obtained was rooted in the 4 N-fixing amino acids (Asp, Glu, Asn, Gln) and 16 triplets of the NAN set. This small, locally phased (commaless) code evidently arose from ambiguous translation on a poly(A) collector strand, in a surface reaction network. Copolymerisation of these amino acids yields polyanionic peptide chains, which could anchor uncharged amide residues to a positively charged mineral surface. From RNA virus structure and replication in vitro, the first genes seemed to be RNA segments spliced into tRNA. Expansion of the code reduced the risk of mutation to an unreadable codon. This step was conditional on initiation at the 5'-codon of a translated sequence. Incorporation of increasingly hydrophobic amino acids accompanied expansion. As codons of the NUN set were assigned most slowly, they received the most nonpolar amino acids. The origin of ferredoxin and Gln synthetase was traced to mid-expansion phase. Surface metabolism ceased by the end of code expansion, as cells bounded by a proteo-phospholipid membrane, with a protoATPase, had emerged. Incorporation of positively charged and aromatic amino acids followed. They entered the post-expansion code by codon capture. Synthesis of efficient enzymes with acid-base catalysis was then possible. Both types of aminoacyl-tRNA synthetases were attributed to this stage. tRNA sequence diversity and error rates in RNA replication indicate the code evolved within 20 million yr in the preIsuan era. These findings on the genetic code provide empirical evidence, from a contemporaneous source, that a surface reaction network, centred on C-fixing autocatalytic cycles, rapidly led to cellular life on Earth.     

Polarity and hydropathic properties of natural amino acids

A new classification of amino acids according to their polarity and symmetric location in the spatial structure of the genetic code is suggested. The polar amino acids are: R, S (codons AGC and AGU), K, N, Q, H, W, C, Y, G, E, D; apolar ones are: T, M, I, P, L, S (codons UCN). Polar and apolar amino acids are grouped into three families whose members possess complementarity with respect to the symmetric structure of the genetic code. Interaction of these complementary polar and apolar amino acids encodes formation of the space structures and ligand-receptor complexes of proteins. Correlation between the polar and hydropathic properties of amino acids is investigated. Normalization of 38 hydrophobicity scales of natural amino acids is carried out. A discrepancy between structures of polar/hydrophilic and apolar/hydrophobic groups of amino acids is demonstrated. According to the signature principle this discrepancy is due to different properties of amino acid side radicals which, in turn, depend on the second component of the reaction and on environmental conditions.     

Is genetic code redundancy related to retention of structural information in both DNA strands?

We have noted that the sense-antisense relationships inherent in the genetic code divide the amino acids into three separate groups. The nature of the amino acids in each group may allow the polypeptides coded by the antisense strand to retain the secondary structure patterns of the translated strand. Also, this relationship requires all but eight of the codons in the eukaryotic code and all but four in the mitochondrial code. Thus, genetic code redundancy could be related to evolutionary pressure toward retention of protein structural information in both strands of DNA.     

Mutations in the alpha8 loop of human APE1 alter binding and cleavage of DNA containing an abasic site

Recent crystallographic studies reveal loops in human AP endonuclease 1 (APE1) that interact with the major and minor grooves of DNA containing apurinic/apyrimidinic (AP) sites. These loops are postulated to stabilize the DNA helix and the flipped out AP residue. The loop alpha8 interacts with the major groove on the 3' side of the AP site. To determine the essentiality of the amino acids that constitute the alpha8 loop, we created a mutant library containing random nucleotides at codons 222-229 that, in wild-type APE1, specify the sequence NPKGNKKN. Upon expression of the library (2 x 10(6) different clones) in Escherichia coli and multiple rounds of selection with the alkylating agent methyl-methane sulfonate (MMS), we obtained approximately 2 x 10(5) active mutants that complemented the MMS sensitivity of AP endonuclease-deficient E. coli. DNA sequencing showed that active mutants tolerated amino acid substitutions at all eight randomized positions. Basic and uncharged polar amino acids together comprised the majority of substitutions, reflecting the positively charged, polar character of the wild-type loop. Asn-222, Asn-226, and Asn-229 exhibited the least mutability, consistent with x-ray data showing that each asparagine contacts a DNA phosphate. Substitutions at residues 226-229, located nearer to the AP site, that reduced basicity or hydrogen bonding potential, increased Km 2- to 6-fold and decreased AP site binding; substitutions at residues 222-225 exhibited lesser effects. This initial mutational analysis of the alpha8 loop supports and extends the conclusion of crystallographic studies that the loop is important for binding of AP.DNA and AP site incision.