Working together or apart : Separating multimodular PKS enzymes into their respective monomodules by replacing the natural intraprotein linkers (illustrated in red in the figure) with a matched docking domain pair from a heterologous PKS system, leads to only small losses in overall in vivo polyketide product and increased efficiency at utilizing polyketide pathway intermediates to prime the biosynthetic process.
Last at last : The terminal step of the gilvocarcin V (GV) biosynthetic pathway is an unusual lactone formation. Here we show that the enzyme, GilR, dehydrogenates the hemiacetal moiety of pregilvocarcin V to the lactone found in GV by using covalently bound FAD.
By miscounting the number of malonyl‐CoA condensations, the stilbene synthase (STS) from Pinus sylvestris forms the previously unknown pentaketide, 2‐malonylresveratrol, in addition to the expected tetraketide resveratrol (see scheme). This is the first time that such tetra‐ and pentaketide‐CoA derivatives have been observed; this suggests that these products might be free intermediates in the biosynthesis of stilbenes.
Flexibility required : We designed intramolecular bipartite tetracysteine sites in loops of p53 and the β‐sheets of EmGFP. We found that ReAsH binding preferentially favors tetracysteine sites with flexible geometries such as loops; flexibility was assessed by comparing Cα B‐factor values. This information is important for directing successful bipartite tetracysteine site designs.
Interactions between C34 and N36 : Synthetic peptides with D ‐amino acid substitutions that mimic the human immunodeficiency virus (HIV) gp41 HR2 region may lead to new peptidic anti‐HIV‐1 drugs that retain potent antiviral activity while being more resistant to proteolytic degradation.
Demystifying Taxol : The structurally complex natural product paclitaxel is among the most important antitumor drugs presently in the clinic. While the mechanism of action is known, the biologically active conformation has been much debated. Herein we summarize the considerable research efforts that have gone into elucidating the bioactive conformation of paclitaxel.
Catch a tiger by the tail : We have demonstrated that by feeding nonmotile mutant C. jejuni bacteria with a neutral azide‐labelled pseudaminic acid precursor we can restore their ability to generate functional flagella. The presence of azido‐pseudaminic acid on the surface of the flagella provides a bio‐orthogonal chemical handle that can be used to modify the flagellar proteins.
Reducing virulence : RhlI catalyzes the synthesis of N‐butanoyl homoserine lactone (BHL), with a minor product N‐hexanoyl homoserine lactone (HHL). By using directed evolution and a genetic screen, RhlI has been engineered for enhanced production of both BHL and HHL at a similar level.
Monooxygenase mutants : A minimal and highly enriched CYP102A1 mutant library was constructed by combining five hydrophobic amino acids in two positions. The library was screened with four different terpene substrates. Eleven variants demonstrated either a strong shift or improved regio‐ or stereoselectivity during oxidation of at least one substrate as compared to CYP102A1 wild type.
Logical, responsible, practical . Enzymatic logic gates that process chemical input signals were used to trigger the release of redox‐active iron ions, which produce reactive oxygen species in a catalytic cascade, and thus result in oxidative damage in biomolecules. Functional coupling between enzymatic logic gates and oxidative damage systems resulted in “smart” biochemical ensembles that are activated upon receiving a certain pattern of biochemical signals.
Sweet medicine : Multimeric glycoconjugates with valencies ranging from one to four were synthesized by click chemistry. Unprecedented adhesion inhibitions of piliated E. coli to human bladder cells were recorded with the multimers; a tetravalent derivative showed inhibitory concentrations 6000‐ and 64‐fold lower than mannose and heptyl α‐D ‐mannoside, respectively.
H 3 R inverse agonists based on an aminopropoxy‐phenyloxazoline framework constitute highly valuable druglike lead compounds that display efficacy in a mouse model of recognition memory.
One enzyme, many substrates . The substrate specificity of a lantibiotic biosynthetic enzyme, lacticin 481 synthetase, was probed by using synthetic prepeptides containing a variety of nonproteinogenic amino acids, including unnatural α‐amino acids, β‐amino acids, D ‐amino acids, and peptoids.
Long‐lasting sweet proteins : The chemoenzymatic synthesis of a triazole (T)‐linked glycosylated C34 fragment from HIV‐1 gp41 is described. The glycopeptide shows high solubility, excellent fusion inhibition, and as shown in the graph, promising protease resistance.
Uranyl ion‐specific DNAzyme : A DNAzyme (lower strand) cleaves the substrate (upper strand) in the presence of the uranyl ion. The enzyme folds into a bulged three‐way‐junction structure with catalytically important nucleotides residing in the bulge. A highly conserved G?A mismatch is also crucial for the enzyme's activity.
Artificial synapses for femtomolar detection : Amperometry at platinized carbon fibre electrodes has been used to unravel the complexity of β‐lapachone's effects on cellular oxidative stress. α‐Lapachone, the pharmacologically inactive para‐quinone isomer, did not display such characteristics, but over longer incubation periods both quinones induced apoptosis. The observed effects were interpreted in terms of two mechanisms involving opposite reactivities of quinones in living cells.
Happy tree‐like friends . Using trypsin and α‐chymotrypsin cleavage sites as models, we show that the protease reactivity of peptide dendrimers, such as the structure illustrated in the figure, can be controlled by the degree of branching. Such a control provides a novel possibility to tune the biological properties of peptide dendrimers, and should be generally useful for their employment as functional biomolecule analogues, for example, in drug delivery applications.
On and off in a flash : We describe a novel, ultrafast, reversible, interstrand RNA photocrosslinking reaction via 3‐cyanovinylcarbazole nucleoside. The interstrand RNA‐photocrosslinking reaction showed a high degree of sequence specificity and can be used in the selection of a target RNA sequence.
A panel of new potential Ras ligands was generated by decorating a tricyclic levoglucosenone‐derived scaffold with aromatic moieties. Some members of the panel show in vitro inhibitory activity toward the nucleotide exchange process on Ras and are toxic to some human cancer cell lines.
Less is more : By starting with a high‐affinity HER2‐binding 3‐helix affibody molecule, we successfully developed 2‐helix small protein binders with 5 nM affinities by using a combination of several different strategies. Our efforts clearly suggest that 2‐helix small proteins against important tumor targets can be obtained by rational protein design and engineering.
