The specificity of peptide chain extension by N-carboxyanhydrides.

We have used amino acids activated by carbonyldiimidazole to study the enantiospecificity of peptide elongation in aqueous solution. Peptide 'primers' Glu10 and Ala3Glu10 were elongated with the enantiomers of arginine, glutamic acid, asparagine, phenylalanine, serine and valine. The homochiral addition was always the more efficient reaction; the enantiospecificity was large in some cases but very small in others. In every case Ala3Glu10 was elongated more efficiently than Glu10.

Inhibition of oligo(glutamine) precipitation by glutamine-containing peptides.

When a solution of glutamine is added to solid 1,1'-carbonyldiimidazole, a mixture of oligo(glutamine)s up to about the 11-mer is formed rapidly. On standing overnight, the solution deposits a precipitate that does not easily redissolve. We have studied the inhibition of this precipitation by glutamine-containing peptides. We find that the alternating peptides (arg.gln)(4) and (arg.gln)(8) are efficient inhibitors of precipitation while arg(5), (glu.gln)(4), and a nonalternating octapeptide of the same composition as (arg.gln)(4) do not inhibit precipitation even though all the arg-containing peptides readily adsorb to oligo(glutamine) precipitates. A possible structural basis for this difference is discussed.

The synthesis of beta-peptides containing guanidino groups.

The synthesis of the beta-peptide 1 by the postsynthetic modification of the corresponding amino-containing peptide 3 is described. The potential of 1 to act as a template for the ligation of complementary negatively-charged peptides is discussed.

Self-organizing biochemical cycles.

I examine the plausibility of theories that postulate the development of complex chemical organization without requiring the replication of genetic polymers such as RNA. One conclusion is that theories that involve the organization of complex, small-molecule metabolic cycles such as the reductive citric acid cycle on mineral surfaces make unreasonable assumptions about the catalytic properties of minerals and the ability of minerals to organize sequences of disparate reactions. Another conclusion is that data in the Beilstein Handbook of Organic Chemistry that have been claimed to support the hypothesis that the reductive citric acid cycle originated as a self-organized cycle can more plausibly be interpreted in a different way.

Oligomerization of alpha-thioglutamic acid.

A decaglutamic acid primer is extended very slowly by alpha-thioglutamic acid. The addition of bicarbonate to the reaction mixture greatly accelerates the reaction. We believe the N-carboxyanhydride of glutamic acid is an intermediate in the accelerated reaction. When K3Fe(CN)6 and bicarbonate are both present in the reaction mixture, oligoglutamic acids up to at least the 15-mer are formed rapidly. The acylating agent is the oxidation product of thioglutamic acid, a diacyldisulfide.

A convenient synthesis of a novel nucleoside analogue: 4-(alpha-diformyl-methyl)-1-(beta-D-ribofuranosyl)-2-pyrimidinone.

The nucleoside analogue 4-(alpha-diformyl-methyl)-1-(beta-D-ribofuranosyl)-2-pyrimidinone (5) was prepared from the corresponding 4-methyl pyrimidinone nucleoside by means of the Vilsmeier reaction. The unprotected nucleoside can be phosphorylated directly with phosphorus oxychloride in triethyl phosphate.

Polyphosphorylation and non-enzymatic template-directed ligation of oligonucleotides.

Oligonucleotide 5'-polyphosphates are formed under potentially prebiotic conditions from oligonucleotide 5'-phosphates and sodium trimetaphosphate. Oligonucleotides activated as polyphosphates undergo template-directed ligation. We believe that these reactions could have produced longer oligonucleotide products from shorter substrates under prebiotic conditions.

Nonenzymatic template-directed reactions on altritol oligomers, preorganized analogues of oligonucleotides.

Altritol nucleic acids (ANAs) are RNA analogues with a phosphorylated D-altritol backbone. The nucleobase is attached at the 2-(S)-position of the carbohydrate moiety. We report that ANA oligomers are superior to the corresponding DNA, RNA, and HNA (hexitol nucleic acid) in supporting efficient nonenzymatic template-directed synthesis of complementary RNAs from nucleoside-5'-phosphoro-2-methyl imidazolides. Activated ANA and HNA monomers do not oligomerize efficiently on DNA, RNA, HNA, or ANA templates.

Origin of life. A simpler nucleic acid.

What was the genetic material of the earliest life forms on Earth if it was not RNA? As Orgel explains in his Perspective, the answer may be simpler nucleic acid polymers perhaps like the RNA analogs called (L)-a-threofuranosyl oligonucleotides or TNAs (Schöning et al.). These molecules have threose rather than ribose in their sugar-phosphate backbones and yet retain many of the properties of RNA including the ability to pair up in double helices.

A comparison of RNA with DNA in template-directed synthesis.

Nonenzymatic template-directed copying of RNA sequences rich in cytidylic acid using nucleoside 5'-(2-methylimidazol-1-yl phosphates) as substrates is substantially more efficient than the copying of corresponding DNA sequences. However, many sequences cannot be copied, and the prospect of replication in this system is remote, even for RNA. Surprisingly, wobble-pairing leads to much more efficient incorporation of G opposite U on RNA templates than of G opposite T on DNA templates.

Nucleic acid duplexes incorporating a dissociable covalent base pair.

We have used molecular modeling techniques to design a dissociable covalently bonded base pair that can replace a Watson-Crick base pair in a nucleic acid with minimal distortion of the structure of the double helix. We introduced this base pair into a potential precursor of a nucleic acid double helix by chemical synthesis and have demonstrated efficient nonenzymatic template-directed ligation of the free hydroxyl groups of the base pair with appropriate short oligonucleotides. The nonenzymatic ligation reactions, which are characteristic of base paired nucleic acid structures, are abolished when the covalent base pair is reduced and becomes noncoplanar. This suggests that the covalent base pair linking the two strands in the duplex is compatible with a minimally distorted nucleic acid double-helical structure.

Peptide-formation on cysteine-containing peptide scaffolds.

Monomeric cysteine residues attached to cysteine-containing peptides by disulfide bonds can be activated by carbonyldiimidazole. If two monomeric cysteine residues, attached to a 'scaffold' peptide Gly-Cys-Glyn-Cys-Glu10, (n = 0, 1, 2, 3) are activated, they react to form the dipeptide Cys-Cys. in 25-65% yield. Similarly, the activation of a cysteine residue attached to the 'scaffold' peptide Gly-Cys-Gly-Glu10 in the presence of Arg5 leads to the formation of Cys-Arg5 in 50% yield. The significance of these results for prebiotic chemistry is discussed.

Oligomerization of L-gamma-carboxyglutamic acid.

Unlike glutamic acid, L-gamma-carboxyglutamic acid does not oligomerize efficiently when treated with carbonyldiimidazole in aqueous solution. However, divalent ions such as Mg2+ catalyze the reaction, and lead to the formation of oligomers in good yield. In the presence of hydroxylapatite, L-gamma-carboxyglutamic acid oligomerizes efficiently in a reaction that proceeds in the absence of divalent ions but is further catalyzed when they are present. After 'feeding' 50 times with activated amino acid in the presence of the Mg2+ ion, oligomers longer than the 20-mer could be detected. The effect of hydroxylapatite on peptide elongation is very sensitive to the nature of the activated amino acid and the acceptor peptide. Glutamic acid oligomerizes more efficiently than L-gamma-carboxyglutamic acid on hydroxylapatite and adds more efficiently to decaglutamic acid in solution. One might, therefore, expect that glutamic acid would add more efficiently than L-gamma-carboxyglutamic acid to decaglutamic acid on hydroxylapatite. The contrary is true--the addition of L-gamma-carboxyglutamic acid is substantially more efficient. This suggests that oligomerization on the surface of hydroxylapatite depends on the detailed match between the structure of the surface of the mineral and the structure of the oligomer.

Nonenzymatic oligomerization reactions on templates containing inosinic acid or diaminopurine nucleotide residues.

The template-directed oligomerization of nucleoside-5'-phosphoro-2-methyl imidazolides on standard oligonucleotide templates has been studied extensively. Here, we describe experiments with templates in which inosinic acid (I) is substituted for guanylic acid, or 2,6-diaminopurine nucleotide (D) for adenylic acid. We find that the substitution of I for G in a template is strongly inhibitory and prevents any incorporation of C into internal positions in the oligomeric products of the reaction. The substitution of D for A, on the contrary, leads to increased incorporation of U into the products. We found no evidence for the template-directed facilitation of oligomerization of A or I through A-I base pairing. The significance of these results for prebiotic chemistry is discussed.

Nonenzymatic synthesis of RNA and DNA oligomers on hexitol nucleic acid templates: the importance of the A structure.

Hexitol nucleic acid (HNA) is an analogue of DNA containing the standard nucleoside bases, but with a phosphorylated 1,5-anhydrohexitol backbone. HNA oligomers form duplexes having the nucleic acid A structure with complementary DNA or RNA oligomers. The HNA decacytidylate oligomer is an efficient template for the oligomerization of the 5'-phosphoroimidazolides of guanosine or deoxyguanosine. Comparison of the oligomerization efficiencies on HNA, RNA, and DNA decacytidylate templates under various conditions suggests strongly that only nucleic acid double helices with the A structure support efficient template-directed synthesis when 5'-phosphoroimidazolides of nucleosides are used as substrates.

Oligomerization of deoxyguanosine 5'-phosphoro-2-methylimidazolide on a polycytidylate template.

The oligomerization of deoxyguanosine 5'-phosphoro-2-methylimidazolide on a polycytidylate template is much less efficient than the oligomerization of the corresponding activated ribonucleotide. Nonetheless oligomers containing up to eight nucleotide residues are detected. The products are 3'-5'-linked oligodeoxyribonucleotides capped at the 5'-terminus with a pyrophosphate-linked monomer.