Polymerization of vinyl fluoride initiated by the system silver nitrate–tetraethyl lead

The initiator system AgNO₃–tetraethyl lead was used for the polymerization of vinyl fluoride. Dimethyl sulfoxide was found to have a specific effect on the half-life of the initiating system, which leads to long reaction times and high conversions at ambient temperatures. This effect was explained by the complexing power of DMSO on silver ions, which slows down the rate of formation of AgEt and permits the polymerization to proceed for longer periods than in other solvents. The kinetics of the polymerization agreed with the scheme developed earlier. Tetraethyl lead acts as a powerful chain transfer agent and determines to a large extent the resulting polymers.     

Prechemistry barriers and checkpoints do not contribute to fidelity and catalysis as long as they are not rate limiting

In the preceding article, “Perspective: Pre-chemistry conformational changes in DNA polymerase mechanisms” contributed by Schlick and coworkers as well as previous studies of these workers (Schlick et al. in Theor Chem Acc 131:1287, 2012; Radhakrishnan and Schlick in J Am Chem Soc 127:13245–13252, 2005; Radhakrishnan and Schlick in Biochem Biophys Res Commun 350:521–529, 2006; Radhakrishnan et al. in Biochemistry 45:15142–15156, 2006; Radhakrishnan and Schlick in Proc Natl Acad Sci USA 101:5970–5975, 2004) have argued that the conformational changes preceding the chemical step contribute to DNA synthesis and to the fidelity of DNA polymerases. In one of our previous investigations (Ram Prasad and Warshel in Proteins 79:2900–2919, 2011), we argued and showed that as long as the free energy barriers associated with any of the prechemistry steps are not rate limiting, they could not contribute to the catalysis and then to the fidelity. Though all our arguments are based on exact and well-defined scientific logics, Schlick and coworkers seem to overlook some of the clear conditions in these arguments and in particular the requirement that the chemical step is rate limiting in their arguments that the prechemistry barriers contribute to the catalysis. In fact, as long as the prechemistry steps are not rate limiting, we have shown that the enzymes cannot carry the memory of the previous steps. We also address other potential misunderstandings about several key issues; First, we clarify that it is misleading to relate the prechemistry proposal to the clear fact that the substrate-induced conformational changes determine the final preorganization (the issue is the height of the barrier of the enzyme substrate system and not the trivial fact that the enzyme has to change its structure when the substrate binds). Second, we address the presumed role of dynamical effects in enzyme catalysis and the assumption that any observable should be explored in studies of biological function even if they are not relevant to the given effect. Third, we clarify that the fidelity cannot be explained or quantified by invoking the induced fit or conformational selection effects but by evaluating the free energy contributions to the rate-limiting steps from the structures of the corresponding systems (that of course can reflect the induce fit structural changes). Overall, we put a major emphasis on clarifying what is the prechemistry proposal and thus on trying to force the reader to focus on the only real controversy. We of course dismiss any implication that our studies cannot explore mutational effects as we actually pioneered such computational studies and we clarify that in studies of chemical rates, the focus must be placed on evaluating the chemical barriers, rather than on irrelevant factors, but that the calculations of the chemical barriers must consider all the factors that determine this barrier (including metal ions) and also examine if needed different problematic proposals such as dynamical effects, tunneling, and prechemistry.     

Ranking asymmetric auctions

We compare the expected revenue in first- and second-price auctions with asymmetric bidders. We consider “close to uniform” distributions with identical supports and show that in the case of identical supports the expected revenue in second-price auctions may exceed that in first-price auctions. We also show that asymmetry over lower valuations has a stronger negative impact on the expected revenue in first-price auctions than in second-price auctions. However, asymmetry over high valuations always increases the revenue in first-price auctions.     

Use of 18O water and ESI-MS detection in subsite characterisation and investigation of the hydrolytic action of an endoglucanase

We present a novel method for investigating subsite-substrate interactions of glycoside hydrolases and the determination of the oligosaccharide cleavage point based on the analysis of the hydrolysis products produced in the presence of ¹⁸O-labelled water. Conventional techniques for such determination of the hydrolysis pattern call for the chemical modification of the substrate, whereas the method presented makes it possible to use natural substrates, utilising the selectivity and sensitivity of mass spectrometry. This method is very useful for the detection and analysis of enzyme-catalysed hydrolysis, provided that the conditions are chosen where ¹⁸O incorporation without the presence of the enzyme is absent or undetectable. Such conditions were found and used in incubations of cellopentaose with the well-characterised endoglucanase Cel5A from Bacillus agaradhaerens. We were able to confirm that the preferred glycoside bond to be hydrolysed is the third one counting from the non-reducing end of the cellopentaose. Thus, cellopentaose prefers to bind from the –3 to the +2 subsites, which is in accordance with published crystallographic data. The main advantage of the method presented is that there is no need for a priori chemical modification/labelling of oligosaccharide substrates, which are processes that can disturb the enzyme–substrate interaction. From ¹⁸O incorporation we could demonstrate that the enzyme also has an oxygen-exchange activity on cellotriose and cellobiose. This is in agreement with the mechanism for transglycosylation and indicates that it is possible for the enzyme to perform such reactions.     

On the numerical quadrature of highly-oscillating integrals I: Fourier transforms

Highly-oscillatory integrals are allegedly difficult to calculate.The main assertion of this paper is that that impression isincorrect. As long as appropriate quadrature methods are used,their accuracy increases when oscillation becomes faster andsuitable choice of quadrature points renders this welcome phenomenonmore pronounced. We focus our analysis on Filon-type quadratureand analyse its behaviour in a range of frequency regimes forintegrals of the form ₀^h f(x)e^{i x} w(x)d x, where h>0 issmall and | | large. Our analysis is applied to modified Magnus methods for highly-oscillatoryordinary differential equations. Received 6 June 2003. Revised 14 October 2003.     

Liquid chromatography-mass spectrometry analysis of enzyme-hydrolysed carboxymethylcellulose for investigation of enzyme selectivity and substituent pattern

A series of celloendoglucanases: Bacillus agaradhaerens Cel 5a, Humicola insolens Cel 5a, H. insolens Cel 7b, H. insolens Cel 45a, Trichoderma reesei Cel 7b, and T. reesei Cel 45a were used to hydrolyse carboxymethylcellulose (CMC) and the hydrolysis products were investigated with a novel liquid chromatography-mass spectrometry (LC-MS) method. Separation was achieved using a graphitised carbon chromatographic column which allowed the use of electrospay compatible eluents. Analysis of the compounds produced during enzyme hydrolysis of CMC is used to understand enzyme selectivities and substitution pattern of CMC. Conventional high-performance anion-exchange chromatography (HPAEC)-pulsed amperometric detection (PAD), size-exclusion chromatography (SEC)-refractive index (RI) detection, and reducing end analysis are also used to analyse enzyme-hydrolysed CMC. The LC-MS method presented allows for a more detailed investigation of hydrolysis products, which facilitates characterisation of both enzymes and substrates.     

The forbidden (J → J + 1) spectrum of CH4 in the ground vibronic state

The pure rotational R-branch spectrum of CH₄ arising from the centrifugal distortion moment has been studied using a simple 12.10-m light-pipe cell and a conventional interferometer. Ten forbidden (J → J + 1) transitions for J = 7 to J = 16 have been observed in the spectral region 80–200 cm^?1 with a theoretical resolution of 0.5 cm^?1. The integrated intensity of the six strongest lines has been measured and was found to be of the order of twice that calculated from the distortion moment obtained earlier from a molecular beam study of the (J = 2) rotational level. In the approximation that frequency shifts due to this excess intensity are neglible, it has been determined that the rotational constant B₀ = (5.245 ± 0.004) cm^?1 and the scalar distortion constant D_S = (1.19 ± 0.09) × 10^?4 cm^?1. It is argued that the excess intensity is due to higher-order terms in the dipole moment operator and the validity of the frequency analysis is considered in this context.     

Through the channel and around the channel: Validating and comparing microscopic approaches for the evaluation of free energy profiles for ion penetration through ion channels

Microscopic calculations of free energy profiles for ion transport through biological ion channels present a very serious challenge to modern simulation approaches. The main problem is due to the major convergence problems associated with the heterogeneous landscape of the electrostatic environment in ion channels and with the need to evaluate the profile associated with the transfer of the ion from bulk water to the channel environment. This problem is compounded by the lack of reliable and relevant benchmarks that can discriminate between alternative approaches. The present study is aimed at reducing the above problems by defining benchmarks that are directly relevant to ion channels and can also give converging results. This is done by constructing a series of models of a truncated gramicidin channel with different numbers of water molecules and by comparing the profiles for going around the channel and through the channel. These discriminating models are then used to validate and compare the adiabatic charging free energy perturbation (FEP) approach combined with an umbrella sampling approach (Warshel, A. J. Phys. Chem. 1982, 86, 2218) and the potential of mean force (PMF) approach used frequently in studies of ion channels. It is found that both approaches work quite well until one moves to the case of the fully solvated channel. In this limit, the PMF approach may give different results for the overall work of going through the channel and around the channel, while the FEP approach gives physically consistent results. The present benchmark also indicates that the weighted histogram analysis method (WHAM) approach does not offer a significant advantage over earlier approaches at least as much as studies of ion channels are concerned. Finally, it is concluded that the FEP approach may be more useful in evaluating the overall barrier for moving ions from water to ion channels and that in some cases it might be beneficial to use the FEP approach for selective points along the channel and then to connect these points by PMF calculations.