Rings, radicals, and regeneration: the early years of a bioorganic laboratory

Abstract

This Perspective provides an overview of the progress in two of the original programs in my research group focused on the biosynthesis of the antibiotics nisin, lacticin 481, fosfomycin, and bialaphos. The path from start-up funds to tenure and beyond offers insights into the opportunities realized and missed along the road.

Figure 1

Structures of the lantibiotics nisin A and lacticin 481, the phosphonate antibiotic fosfomycin, and the phosphinate phosphinothricin. Lanthionine (Lan) residues in the lantibiotics are shown in red and methyllanthionine (MeLan) residues in blue. The two methyl groups in fosfomycin and phosphinothricin that are considered to be derived from methylcobalamin are highlighted in green.

Figure 2

Structures of vitamin B₁₂ (X = cyanide), and the natural cofactors methylcobalamin (X = CH₃) and adenosylcobalamin (X = 5′-deoxyadenosine).

Figure 3

The post-translational maturation process of nisin as an example of a class I lantibiotic. The prepeptide NisA is ribosomally synthesized, followed by NisB catalyzed dehydration of underlined Ser and Thr residues of NisA. NisC catalyzes the conjugate addition of Cys residues in a regio- and stereoselective manner to five of the Dha (green) and Dhb (magenta) residues to generate five cyclic thioethers: one lanthionine (red) and four methyllanthionines (blue). After dehydration/cyclization is complete, the unmodified leader peptide is proteolytically removed by the protease NisP. The sequence of the leader peptide is MSTKDFNLDLVSVSKKDSGASPR. Abu, S-2-aminobutyric acid.

Figure 4

The post-translational maturation process of lacticin 481, a class II lantibiotic. LctM catalyzes the anti elimination of water from the underlined Ser and Thr residues in the propeptide region of LctA to generate Dha and Dhb residues. LctM also catalyzes the conjugate addition of three Cys residues in a regioselective manner to three of the dehydrated residues to generate three cyclic thioethers, one methyllanthionine (blue) and two lanthionines (red). The leader peptide is proteolytically removed by the N-terminal protease domain of LctT that also excretes the final product. The sequence of the leader peptide is MKEQNSFNLLQEVTESELDLILGA. Abu, S-2-aminobutyric acid.

Figure 5

Various post-translational modifications of proteinogenic amino acids found in lantibiotic family members. The stereochemistry of the dihydroxyproline is currently unknown. Thioviramide contains an aminovinylcysteine suggesting it may be a lantibiotic, but it has not yet been shown to be gene encoded. It also contains the thioamides shown in the figure.

Figure 6

Conventional and unusual mechanisms, of B₁₂-dependent methyl transfer. The corrin ligand ring system (Figure 2) is schematically represented by a square. L may be the benzimidazole ligand of B₁₂ or an amino acid ligand from the protein.

Scheme 1

Preparation of dehydropeptides by oxidative elimination.

Scheme 2

Attempted biomimetic synthesis of the A and B-rings of nisin.

Scheme 3

Conjugate addition of external thiol nucleophiles to Dha.

Scheme 4

Stereoselective synthesis of thioglycopeptides.,

Scheme 5

NAD⁺-dependent oxidation of phosphite by PTDH.

Scheme 6

Use of PTDH for cofactor regeneration.

Scheme 7

Proposed mechanism of enzymatic Lan and MeLan formation.

Scheme 8

Several members of the phosphonate/phosphinate class of natural products. The phosphinothricin moiety in bialaphos is highlighted in red.

Scheme 9

Proposed biosynthetic pathways of fosfomycin and bialaphos.

Scheme 10

Possible heterolytic mechanism of P-methyltransferase (Pmet).

Scheme 11

Possible homolytic mechanism of P-methyltransferase (Pmet).

Scheme 12

Revised biosynthetic pathway of fosfomycin

Scheme 13

Labeling studies on fosfomycin biosynthesis.

Scheme 14

Proposed mechanism for Fom3.