Probable Role of Clavaminic Acid as the Terminal Intermediate in the Common Pathway to Clavulanic Acid and the Antipodal Clavam Metabolites

Abstract

Emerging chemical and genetic evidence suggests that separate biochemical solutions have evolved to synthesize the four known classes of β-lactam antibiotics. One of these classes contains clavulanic acid (1) and a family of structurally related but antipodal clavam metabolites 2−5, 7, and 8 which lack a carboxylate at C-3, have a different oxidation state, and exhibit stereochemical features at C-2. Previous work has demonstrated the incorporation of ornithine/arginine in the identical regiochemical sense in all of these natural products, and has established the common intermediacy of the monocyclic β-lactam proclavaminic acid (12) as well. In this paper the quite advanced bicyclic intermediate clavaminic acid (14) has been synthesized in doubly 13C-labeled form by preparative incubation of recombinant clavaminate synthase. The intact and equally efficient incorporation of 14 into valclavam (7) and 2-(2-hydroxyethyl)clavam (8), together with 18O2-incorporation experiments, has been interpreted to define clavaminic acid as the final intermediate shared in the biosynthesis of clavulanic acid and the antipodal clavams. A mechanistic rationale of this interrelationship and the late stages of the respective biosyntheses is proposed.

ORF17 from the Clavulanic Acid Biosynthesis Gene Cluster Catalyzes the ATP-dependent Formation of N-Glycyl-clavaminic Acid

“…Steps in the pathway leading to (3S,5S)-clavaminic acid, the proposed branch point between the biosynthesis of clavulanic acid and the (3S,5S)-clavams (13), have been characterized (Fig. 1b), and the enzymes that catalyze them have been identified.…”

The Paralogous Pairs of Genes Involved in Clavulanic Acid and Clavam Metabolite Biosynthesis Are Differently Regulated in Streptomyces clavuligerus

“…1B) (10). However, in S. clavuligerus, the genes involved in the biosynthesis of clavulanic acid and the 5S clavams reside in three distinct gene clusters that are not physically linked (K. Tahlan, H. U.…”

“…Next, in a series of two sequential reactions, CAS converts proclavaminic acid to clavaminic acid (4,36). Clavaminic acid is thought to be the branch point of the pathway leading to clavulanic acid and the 5S clavams (10). The only other step known in the pathway beyond clavaminic acid is the reduction of clavaldehyde to clavulanic acid by the action of the enzyme clavulanic acid dehydrogenase (27), which is encoded by cad, a late gene from the clavulanic acid gene cluster (15).…”

Carboxyethylarginine Synthase Genes Show Complex Cross-Regulation in Streptomyces clavuligerus

“…Due to its clinical value (1), the biochemical and genetic aspects of clavulanic acid biosynthesis in S. clavuligerus have been subjected to intensive research in recent years. It is now known that the biosynthetic pathway to clavulanic acid is partly shared with the pathway for structurally related 5S clavam metabolites that are also produced by S. clavuligerus, up to the level of clavaminic acid (2). The pathway branches at this point and goes on to form either clavulanic acid or 5S clavams (Fig.…”