Lewis acid mediated hydrosilylation of alkynes and alkenes on non-oxidized hydride-terminated porous silicon derivatizes the surface with alkenyl and alkyl functionalities, respectively. A very broad range of chemical groups may be incorporated, allowing for tailoring of the interfacial characteristics of the material. The reaction is shown to protect and stabilize porous silicon surfaces from atmospheric or direct chemical attack without compromising its valuable material properties such as high porosity, surface area and visible room-temperature photoluminescence. The reaction is shown to act on alkenes and alkynes of all possible regiochemistries (terminal and internal alkynes; mono-, cis- and trans-, di-, tri-, and tetrasubstituted alkenes). FTIR as well as liquid- and solid-state NMR spectroscopies show anti-Markovnikov addition and cis stereochemistry in the case of hydrosilylated terminal alkynes. Material hydrosilylated with long-chain hydrophobic alkynes and alkenes shows a substantially slower surface oxidation and hydrolysis rate in air as monitored by long-term FTIR monitoring and chemography. BJH and BET measurements reveal that the surface area and average pore size of the material are reduced only slightly after hydrosilylation, indicating that the porous silicon skeleton remains intact. Elemental analysis and SIMS depth profiling show a consistent level of carbon incorporation throughout the porous silicon which demonstrates that the reaction occurs uniformly throughout the depth of the film. The effects of functionalization on photoluminescence were investigated and are shown to depend on the organic substituents.

The chemical diversity of natural products is fueled by the emergence and ongoing evolution of biosynthetic pathways in secondary metabolism [1][2][3][4][5] . However, co-evolution of enzymes as functional assemblies for metabolic diversification is not well understood, especially at the biochemical level. Here, two parallel enzyme assemblies with an extraordinarily high sequence identity form a β-branched cyclopropane in the curacin A (Cur), and a vinyl chloride group in the jamaicamide (Jam) pathways, respectively. The assemblies include a halogenase (Hal), a 3-hydroxy-3-methylglutaryl (HMG) enzyme cassette for β-branching and an enoyl reductase domain (ER). Bioinformatic analysis indicated that the corresponding genes were inserted into modular polyketide synthases (PKSs) via acyltransferase (AT) domain replacement. The Hal from CurA, and the dehydratases (ECH 1 s) and decarboxylases (ECH 2 s) within the HMG enzyme cassettes and ERs from both Cur and Jam were assessed biochemically to determine the mechanism of cyclopropane and vinyl chloride formation. Unexpectedly, the polyketide β-branching pathway was modified by introduction of a γ-chlorination step on (S)-HMG mediated by Cur Hal, a nonheme Fe II , α-ketoglutarate (α-KG)-dependent halogenase 6 . In a divergent scheme, Cur ECH 2 was found to catalyze formation of the α,β C=C enoyl thioester, whereas Jam ECH 2 formed a vinyl chloride moiety by selectively generating the corresponding β,γ C=C (enoyl thioester) of the 3-methyl-4-chloroglutaconyl decarboxylation product. A non-conserved Tyr 82 residue in Cur ECH 2

Bacterial type I polyketide synthases assemble structurally diverse natural products of significant clinical value from simple metabolic building blocks. The synthesis of these compounds occurs in a processive fashion along a large multi-protein complex. Transfer of the acyl intermediate across inter-polypeptide junctions is mediated, at least in large part, by N-and C-terminal docking domains. We report here a comprehensive analysis of the binding affinity and selectivity for the complete set of discrete docking domain pairs in the pikromycin and erythromycin PKS systems. Despite disconnection from their parent module, each cognate pair of docking domains retained exquisite binding selectivity. Further insights were obtained by X-ray crystallographic analysis of the PikAIII/ PikAIV docking domain interface. This new information revealed a series of key interacting residues that enabled development of a structural model for the recently proposed H2-T2 class of polypeptides involved in PKS intermodular molecular recognition.

An unexpected biochemical strategy for chain initiation is described for the loading module of the polyketide synthase of curacin A, an anticancer lead derived from the marine cyanobacterium Lyngbya majuscula. A central GCN5-related N-acetyltransferase (GNAT) domain bears bifunctional decarboxylase/S-acetyltransferase activity, both unprecedented for the GNAT superfamily. A CurA loading tridomain, consisting of an adaptor domain, the GNAT domain, and an acyl carrier protein, was assessed biochemically, revealing that a domain showing homology to GNAT (GNAT(L)) catalyzes (i) decarboxylation of malonyl-coenzyme A (malonyl-CoA) to acetyl-CoA and (ii) direct S-acetyl transfer from acetyl-CoA to load an adjacent acyl carrier protein domain (ACP(L)). Moreover, the N-terminal adapter domain was shown to facilitate acetyl-group transfer. Crystal structures of GNAT(L) were solved at 1.95 angstroms (ligand-free form) and 2.75 angstroms (acyl-CoA complex), showing distinct substrate tunnels for acyl-CoA and holo-ACP(L) binding. Modeling and site-directed mutagenesis experiments demonstrated that histidine-389 and threonine-355, at the convergence of the CoA and ACP tunnels, participate in malonyl-CoA decarboxylation but not in acetyl-group transfer. Decarboxylation precedes acetyl-group transfer, leading to acetyl-ACP(L) as the key curacin A starter unit.

SUMMARY The mycobacterial biotin protein ligase (MtBPL) globally regulates lipid metabolism in Mtb through the posttranslational biotinylation of acyl coenzyme A carboxylases involved in lipid biosynthesis that catalyze the first step in fatty acid biosynthesis and pyruvate coenzyme A carboxylase, a gluconeogenic enzyme vital for lipid catabolism. Here we describe the design, development and evaluation of a rationally designed bisubstrate inhibitor of MtBPL. This inhibitor displays potent sub-nanomolar enzyme inhibition and antitubercular activity against multi- and extensively drug resistant Mtb strains. We show that the inhibitor decreases in vivo protein biotinylation of key enzymes involved in fatty acid biosynthesis and that the anti-bacterial activity is MtBPL-dependent. Additionally, the gene encoding BPL was found to be essential in M. smegmatis. Finally, the X-ray co-crystal structure of inhibitor bound MtBPL was solved providing detailed insight for further structure-activity analysis. Collectively, these data suggest that MtBPL is a promising target for further antitubercular therapeutic development.