The construction of mirror-image biological systems may open the next frontier for biomedical technology development and discovery. Here we have designed and chemically synthesized a mutant version of the thermostable Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4) consisting of d-amino acids. With a total peptide length of 358 amino acid residues, it is the largest chemically synthesized d-amino acid protein reported to date. We show that the d-polymerase is able to amplify a 120-bp l-DNA sequence coding for the Escherichia coli 5S ribosomal RNA gene rrfB by mirror-image polymerase chain reaction, and that both the natural and mirror-image systems operate with strict chiral specificity. The development of efficient miPCR systems may lead to many practical applications, such as mirror-image systematic evolution of ligands by exponential enrichment for the selection of therapeutically promising nuclease-resistant l-nucleic acid aptamers.

Toward building a chirally inverted version of the central dogma of molecular biology, Zhu and colleagues realized the transcription of a mirror-image gene into L-RNA and reverse transcription into L-DNA by synthetic D-polymerases. They enzymatically transcribed full-length mirror-image 5S rRNA and demonstrated a comprehensive mirror-image transcription, reverse transcription, PCR, and sequencing toolbox.

This paper describes recent advances of one-pot multi-segment condensation strategies based on kinetically controlled strategies and/or protecting group-removal strategies in chemical protein synthesis.