Are Free Radicals Involved in IspH Catalysis? An EPR and Crystallographic Investigation

The [4Fe–4S] protein IspH in the methylerythritol phosphate isoprenoid biosynthesis pathway is an important anti-infective drug target, but its mechanism of action is still the subject of debate. Here, by using electron paramagnetic resonance (EPR) spectroscopy and ²H, ¹⁷O, and ⁵⁷Fe isotopic labeling, we have characterized and assigned two key reaction intermediates in IspH catalysis. The results are consistent with the bioorganometallic mechanism proposed earlier, and the mechanism is proposed to have similarities to that of ferredoxin, thioredoxin reductase, in that one electron is transferred to the [4Fe–4S]²⁺ cluster, which then performs a formal two-electron reduction of its substrate, generating an oxidized high potential iron–sulfur protein (HiPIP)-like intermediate. The two paramagnetic reaction intermediates observed correspond to the two intermediates proposed in the bioorganometallic mechanism: the early π-complex in which the substrate’s 3-CH₂OH group has rotated away from the reduced iron–sulfur cluster, and the next, η³-allyl complex formed after dehydroxylation. No free radical intermediates are observed, and the two paramagnetic intermediates observed do not fit in a Birch reduction-like or ferraoxetane mechanism. Additionally, we show by using EPR spectroscopy and X-ray crystallography that two substrate analogues (<b>4</b> and <b>5</b>) follow the same reaction mechanism

Stereoisomeric Composition of Natural Myrtucommulone A

Myrtucommulone A (MC A) (<b>1</b>), isolated from <i>Myrtus communis</i> (myrtle), shows the same pharmacological activity for inhibition of inflammation and induction of apoptosis as synthetic MC A, which consists of three stereoisomers, i.e., two enantiomers and one meso form. This led to the question of whether the natural MC A is a pure stereoisomer or a mixture of stereoisomers. The specific rotation and electronic circular dichroism (ECD) data of natural MC A (<b>1</b>) as well as of a pentacyclic derivative <b>4</b> revealed that naturally occurring MC A (<b>1</b>) consists of the racemate and the meso form in a 1:1 ratio. A probable precursor of MC A (<b>1</b>), nor-semimyrtucommulone (<b>5</b>), was also isolated from myrtle as a racemate. The absolute configurations of the enantiomers of <b>1</b> and <b>5</b> were determined using a combination of experimental and quantum-chemical calculated ECD spectra

Tetra- and Pentacyclic Triterpene Acids from the Ancient Anti-inflammatory Remedy Frankincense as Inhibitors of Microsomal Prostaglandin E₂ Synthase‑1

The microsomal prostaglandin E₂ synthase (mPGES)-1 is the terminal enzyme in the biosynthesis of prostaglandin (PG)­E₂ from cyclooxygenase (COX)-derived PGH₂. We previously found that mPGES-1 is inhibited by boswellic acids (IC₅₀ = 3–30 μM), which are bioactive triterpene acids present in the anti-inflammatory remedy frankincense. Here we show that besides boswellic acids, additional known triterpene acids (i.e., tircuallic, lupeolic, and roburic acids) isolated from frankincense suppress mPGES-1 with increased potencies. In particular, 3α-acetoxy-8,24-dienetirucallic acid (<b>6</b>) and 3α-acetoxy-7,24-dienetirucallic acid (<b>10</b>) inhibited mPGES-1 activity in a cell-free assay with IC₅₀ = 0.4 μM, each. Structure–activity relationship studies and docking simulations revealed concrete structure-related interactions with mPGES-1 and its cosubstrate glutathione. COX-1 and -2 were hardly affected by the triterpene acids (IC₅₀ > 10 μM). Given the crucial role of mPGES-1 in inflammation and the abundance of highly active triterpene acids in frankincence extracts, our findings provide further evidence of the anti-inflammatory potential of frankincense preparations and reveal novel, potent bioactivities of tirucallic acids, roburic acids, and lupeolic acids