Optical Modulation of Antibiotic Resistance by Photoswitchable Cystobactamids

The rise of antibiotic resistance causes a serious health care problem, and its counterfeit demands novel, innovative concepts. The combination of photopharmacology, enabling a light-controlled reversible modulation of drug activity, with antibiotic drug design has led to first photoswitchable antibiotic compounds derived from established scaffolds. In this study, we converted cystobactamids, gyrase-inhibiting natural products with an oligoaryl scaffold and highly potent antibacterial activities, into photoswitchable agents by inserting azobenzene in the N-terminal part and/or an acylhydrazone moiety near the C-terminus, yielding twenty analogs that contain mono- as well as double-switches. Antibiotic and gyrase inhibition properties could be modulated 3.4-fold and 5-fold by light, respectively. Notably, the sensitivity of photoswitchable cystobactamids towards two known resistance factors, the peptidase AlbD and the scavenger protein AlbA, was light-dependent. While irradiation of an analog with an N-terminal azobenzene with 365 nm light led to less degradation by AlbD, the AlbA-mediated inactivation was induced. This provides a proof-of-principle that resistance towards photoswitchable antibiotics can be optically controlled.     

Modeling the influence of inherent envelope fluctuations in simultaneous masking experiments

Masked thresholds are measured and simulated for bandpass-noise signals ranging in bandwidth from 4 to 256 Hz in the presence of a masking bandpass noise also ranging in bandwidth from 4 to 256 Hz. Signal and masker are centered at 2 kHz. To investigate the role of temporal processing in simultaneous masking, simulations were performed with the modulation-filterbank model by Dau et al. [J. Acoust. Soc Am. 102, 2906-2919 (1997)]. For a fixed masker bandwidth, thresholds are independent of the signal bandwidth as long as the signal bandwidth does not exceed the masker bandwidth and thresholds decrease with increasing masker bandwidth in those conditions. A simple modulation-low-pass filter (energy integrator) would be sufficient to describe the experimental results in those conditions. In contrast, the processing by a modulation filterbank is necessary to account for the conditions of "asymmetry of masking," where thresholds for signals with bandwidths larger than the masker bandwidth are much lower than those for the reversed condition. In those conditions, the modulation-filterbank model is able to use the inherent higher modulation frequencies of the signal as an additional cue.     

Range effects in the identification of lateral position

Experiments on the identification of interaural time and interaural amplitude differences were conducted to evaluate the effects of stimulus range on identification performance. Three stimulus sets, large range (LR), small-range center (SRC), and small-range side (SRS), were used in experiments on interaural time and amplitude identification. As expected, data for both sets of measurements show worse resolution for LR than SRC or SRS, demonstrating that the ability to distinguish between two fixed interaural differences can be strongly influenced by the total range of such differences in the stimulus set.     

In vitro reconstitution of α-pyrone ring formation in myxopyronin biosynthesis

Myxopyronins are α-pyrone antibiotics produced by the terrestrial bacterium Myxococcus fulvus Mx f50 and possess antibacterial activity against Gram-positive and Gram-negative pathogens. They target the bacterial RNA polymerase (RNAP) “switch region” as non-competitive inhibitors and display no cross-resistance to the established RNAP inhibitor rifampicin. Recent analysis of the myxopyronin biosynthetic pathway led to the hypothesis that this secondary metabolite is produced from two separate polyketide parts, which are condensed by the stand-alone ketosynthase MxnB. Using in vitro assays we show that MxnB catalyzes a unique condensation reaction forming the α-pyrone ring of myxopyronins from two activated acyl chains in form of their β-keto intermediates. MxnB is able to accept thioester substrates coupled to either N-acetylcysteamine (NAC) or a specific carrier protein (CP). The turnover rate of MxnB for substrates bound to CP was 12-fold higher than for NAC substrates, demonstrating the importance of protein–protein interactions in polyketide synthase (PKS) systems. The crystal structure of MxnB reveals the enzyme to be an unusual member of the ketosynthase group capable of binding and condensing two long alkyl chains bound to carrier proteins. The geometry of the two binding tunnels supports the biochemical data and allows us to propose an order of reaction, which is supported by the identification of novel myxopyronin congeners in the extract of the producer strain. Insights into the mechanism of this unique condensation reaction do not only expand our knowledge regarding the thiolase enzyme family but also opens up opportunities for PKS bioengineering to achieve directed structural modifications.     

Structure and Biosynthesis of Crocagins: Polycyclic Posttranslationally Modified Ribosomal Peptides from Chondromyces crocatus

Secondary metabolome mining efforts in the myxobacterial multiproducer of natural products, Chondromyces crocatus Cm c5, resulted in the isolation and structure elucidation of crocagins, which are novel polycyclic peptides containing a tetrahydropyrrolo[2,3-b]indole core. The gene cluster was identified through an approach combining genome analysis, targeted gene inactivation in the producer, and in vitro experiments. Based on our findings, we developed a biosynthetic scheme for crocagin biosynthesis. These natural products are formed from the three C-terminal amino acids of a precursor peptide and thus belong to a novel class of ribosomally synthesized and post-translationally modified peptides (RiPPs). We demonstrate that crocagin A binds to the carbon storage regulator protein CsrA, thereby inhibiting the ability of CsrA to bind to its cognate RNA target.     

A Note on normalizations of orbit closures

We give a negative answer to a question by J. M. Landsberg on the nature of normalizations of orbit closures. A counterexample originates from the study of complex, ternary, cubic forms.     

The comparative effects of signal sensation level and sound-pressure level on interaural time discrimination

A series of experiments was performed to examine the extent to which precision of interaural time discrimination depends on the sound-pressure level (SPL) and/or sensation level (SL) of the signal. All experiments used a tone burst signal and a continuous white noise masker, which was either diotic or interaurally phase reversed. Results of the first experiment indicate that (1) at equal signal SLs, interaural time and intensity discrimination is more precise when measured with the added diotic noise, and (2) addition of the phase reversed noise, previously shown to cause less precise interaural time discrimination, has a similar effect on interaural intensity discrimination. In the second experiment, interaural time JNDs for a signal of constant SPL were measured as a function of noise level. Results show that a low-level diotic noise can benefit interaural time discrimination, particularly at 500 Hz. The third and fourth experiments were performed to measure interaural time discrimination as a function of increasing signal SPL but constant signal-to-noise ratio. The data show the JND decreasing with increasing signal SPL at nearly the same rate with or without the added noise, indicating that an increase in signal-to-noise ratio is not necessary for improved discrimination.     

Modeling cochlear dynamics: interrelation between cochlea mechanics and psychoacoustics

A model of the cochlea was used to bridge the gap between model approaches commonly used to investigate phenomena related to otoacoustic emissions and more filter-based model approaches often used in psychoacoustics. In the present study, a nonlinear and active one-dimensional transmission line model was developed that accounts for several aspects of physiological data with a single fixed parameter set. The model shows plausible excitation patterns and an input-output function similar to the linear-compressive-linear function as hypothesized in psychoacoustics. The model shows realistic results in a two-tone suppression paradigm and a plausible growth function of the 2f(1)-f(2) component of distortion product otoacoustic emissions. Finestructure was found in simulated stimulus-frequency otoacoustic emissions (SFOAE) with realistic levels and rapid phase rotation. A plausible "threshold in quiet" including finestructure and spontaneous otoacoustic emissions (SOAE) could be simulated. It is further shown that psychoacoustical data of modulation detection near threshold can be explained by the mechanical dynamics of the modeled healthy cochlea. It is discussed that such a model can be used to investigate the representation of acoustic signals in healthy and impaired cochleae at this early stage of the auditory pathway for both, physiological as well as psychoacoustical paradigms.