Contemporary
chemical protein synthesis has been dramatically advanced
over the past few decades, which has enabled chemists to reach the
landscape of synthetic biomacromolecules. Chemical synthesis can produce
synthetic proteins with precisely controlled structures which are
difficult or impossible to obtain via gene expression systems. Herein,
we summarize the key enabling ligation technologies, major strategic
developments, and some selected representative applications of synthetic
proteins and provide an outlook for future development.
Trimethylsilyl
(TMS) groups present outstanding NMR probes of biological
macromolecules as they produce intense singlets in 1H NMR
spectra near 0 ppm, where few other proton resonances occur. We report
a system for genetic encoding of N
6-(((trimethylsilyl)methoxy)carbonyl)-l-lysine (TMSK) for site-specific incorporation into proteins.
The system is based on pyrrolysyl-tRNA synthetase mutants, which deliver
proteins with high yield and purity in vivo and in
cell-free protein synthesis. As the TMS signal can readily be identified
in 1D 1H NMR spectra of high-molecular weight systems without
the need of isotopic labeling, TMSK delivers an excellent site-specific
NMR probe for the study of protein structure and function, which is
both inexpensive and convenient. We demonstrate the utility of TMSK
to detect ligand binding, measure the rate of conformational change,
and assess protein dimerization by paramagnetic relaxation enhancement.
In addition, we present a system for dual incorporation of two different
unnatural amino acids (TMSK and O-tert-butyl-tyrosine) in the same protein in quantities sufficient for
NMR spectroscopy. Close proximity of the TMS and tert-butyl groups was readily detected by nuclear Overhauser effects.
A PCR-based genetic screening experiment targeting the dTDP-glucose-4,6-dehydratase gene revealed that a marine sediment-derived strain, Streptomyces sp. 7-145, had the potential to produce glycosidic antibiotics. Chemical investigation of culture extracts of this strain yielded two new 6-deoxyhexose-containing antibiotics, 11',12'-dehydroelaiophylin (1) and 11,11'-O-dimethyl-14'-deethyl-14'-methylelaiophylin (2), together with four known elaiophylin analogues (3-6). Their structures were determined by extensive NMR, MS, and CD analyses. Compounds 1, 3, 4, and 6 showed good antibacterial activity against methicillin-resistant Staphylococcus aureus and vancomycin-resistant enterococci pathogens.
Analysis of the whole genome sequence of Streptomyces sp. IMB7-145 revealed the presence of seven type I polyketide synthase biosynthetic gene clusters, one of which was highly homologous to the biosynthetic gene cluster of azalomycin F. Detailed bioinformatic analysis of the modular organization of the PKS gene suggested that this gene is responsible for niphimycin biosynthesis. Guided by genomic analysis, a large-scale cultivation ultimately led to the discovery and characterization of four new niphimycin congeners, namely, niphimycins C-E (1-3) and 17-O-methylniphimycin (4). The configurations of most stereocenters of niphimycins have not been determined to date. In the present study, the relative configurations were elucidated by spectroscopic analysis, including J-based analysis and the CNMR database method. Further, the full absolute configurations of niphimycins were completely proposed for the first time based on biosynthetic gene cluster analysis of the ketoreductase and enoylreductase domains for hydroxy- and methyl-bearing stereocenters. Compounds 1, 3, 4, and niphimycin Iα (5) showed antimicrobial activity against methicillin-resistant Staphylococcus aureus and vancomycin-resistant enterococci (MIC: 8-64 μg/mL), as well as cytotoxicity against the human HeLa cancer cell line (IC: 3.0-9.0 μM). In addition, compounds 1 and 5 displayed significant activity against several Mycobacterium tuberculosis clinical isolates (MIC: 4-32 μg/mL).
The chemical ligation of two unprotected peptides to generate a natural peptidic linkage specifically at the C‐ and N‐termini is a desirable goal in chemical protein synthesis but is challenging because it demands high reactivity and selectivity (chemo‐, regio‐, and stereoselectivity). We report an operationally simple and highly effective chemical peptide ligation involving the ligation of peptides with C‐terminal salicylaldehyde esters to peptides with N‐terminal cysteine/penicillamine. The notable features of this method include its tolerance of steric hinderance from the side groups on either ligating terminus, thereby allowing flexible disconnection at sites that are otherwise difficult to functionalize. In addition, this method can be expanded to selective desulfurization and one‐pot ligation‐desulfurization reactions. The effectiveness of this method was demonstrated by the synthesis of VISTA (216‐311), PD‐1 (192‐288) and Eglin C.
A new photochromic coordination network is constructed via the combination of viologen and carboxylate fragments. Close packing of the two-fold diamond-like interwoven mode creates electron transfer bridges for the acceptor and donor components. Radicals are generated upon photoirradiation and are accompanied by a color change from yellow to blue-green.
Saccharothrixones A-C (1-3), three new aromatic polyketide seco-tetracenomycins, and saccharothrixone D (4), a new tetracenomycin analogue possessing opposite configurations at all of the stereogenic centers, were isolated from the marine-derived actinomycete Saccharothrix sp. 10-10. Compounds 1-3 represent the first examples of seco-tetracenomycins where the quinone ring B is cleaved and re-formed into a furanone ring. Their structures were elucidated by spectroscopic analyses and ECD calculations. The absolute configuration of 4 was confirmed by single-crystal X-ray diffraction analysis. Saccharothrixone D (4) showed in vitro cytotoxic activity against the HepG2 cancer cell line with an IC50 value of 7.5 μM.
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