Enable Bond Formation Research Articles

Interface Formation during Collision Welding of Aluminum

Collision welding is a high-speed joining technology based on the plastic deformation of at least one of the joining partners. During the process, several phenomena like the formation of a so-called jet and a cloud of particles occur and enable bond formation. However, the interaction of these phenomena and how they are influenced by the amount of kinetic energy is still unclear. In this paper, the results of three series of experiments with two different setups to determine the influence of the process parameters on the fundamental phenomena and relevant mechanisms of bond formation are presented. The welding processes are monitored by different methods, like high-speed imaging, photonic Doppler velocimetry and light emission measurements. The weld interfaces are analyzed by ultrasonic investigations, metallographic analyses by optical and scanning electron microscopy, and characterized by tensile shear tests. The results provide detailed information on the influence of the different process parameters on the classical welding window and allow a prediction of the different bond mechanisms. They show that during a single magnetic pulse welding process aluminum both fusion-like and solid-state welding can occur. Furthermore, the findings allow predicting the formation of the weld interface with respect to location and shape as well as its mechanical strength.

Interface Formation during Collision Welding of Aluminum

Collision welding is a high-speed joining technology based on the plastic deformation of at least one of the joining partners. During the process, several phenomena like the formation of a so-called jet and a cloud of particles occur and enable bond formation. However, the interaction of these phenomena and how they are influenced by the amount of kinetic energy is still unclear. In this paper, the results of three series of experiments with two different setups to determine the influence of the process parameters on the fundamental phenomena and relevant mechanisms of bond formation are presented. The welding processes are monitored by different methods, like high-speed imaging, photonic Doppler velocimetry and light emission measurements. The weld interfaces are analyzed by ultrasonic investigations, metallographic analyses by optical and scanning electron microscopy, and characterized by tensile shear tests. The results provide detailed information on the influence of the different process parameters on the classical welding window and allow a prediction of the different bond mechanisms. They show that during a single magnetic pulse welding process aluminum both fusion-like and solid-state welding can occur. Furthermore, the findings allow predicting the formation of the weld interface with respect to location and shape as well as its mechanical strength.

Abstract 686: Drugging an undruggable pocket: The biochemical mechanism of covalent KRASG12C inhibitors

Abstract We describe the biochemical mechanism of the covalent KRASG12C inhibitors ARS-853 and ARS-1620. Activating mutations in KRAS are among the most common mutations found in cancer. The KRASG12C mutation in particular is observed in approximately 15 % of non-small cell lung adenocarcinoma, 3 % of colorectal adenocarcinoma and 1 % of pancreatic adenocarcinoma. Until recently, KRAS had been considered undruggable due to the lack of clearly defined pockets that might support binding of small molecules, and the difficulty of targeting the nucleotide binding site due to the high affinity of GDP and GTP. However several years ago small molecules were discovered that bind an inducible pocket near the switch II region and covalently target the mutated cysteine in KRASG12C, trapping KRASG12C in a nonproductive GDP-bound state. Subsequent optimization of these compounds yielded the recently described inhibitors ARS-853 and ARS-1620, the first compounds that directly inhibit KRAS with high potency in cells and animals. While the biological activity of the inhibitors has been described, the biochemical mechanism of how the compounds achieve potent inhibition remained incompletely understood. We now show through biochemical kinetics studies that the activity of ARS-853 and ARS-1620 is primarily driven by KRAS-mediated catalysis of the chemical step of covalent bond formation with cysteine 12 in KRASG12C, rather than by high reversible binding affinity. The reversible inhibition constant (Ki) for both ARS-853 and ARS-1620 is well above the highest compound concentration tested (64 µM, to avoid solubility limitations), likely in the hundreds of micromolar range, while the rate of the chemical step (kinact) is fast. We confirm by several independent means that there is no detectable reversible binding affinity of the inhibitors for KRAS up to at least 32 µM, and show that the rapid chemical reaction is not due to high inherent reactivity of cysteine 12 in KRAS, nor to high intrinsic reactivity of the inhibitors. The results imply that the inhibitors do bind reversibly to KRAS to enable bond formation, but that binding is weak and primarily serves to orient the electrophile. The KRAS-dependent activation of covalent bond formation of ARS-853 and ARS-1620 with the mutated cysteine 12 is reminiscent of mechanism-based or suicide covalent enzyme inhibition, and explains both the high selectivity of the inhibitors for this cysteine relative to other cellular cysteines, and their potent overall activity despite exhibiting poor reversible affinity. The mechanism described here therefore resolves how an induced, shallow and dynamic pocket that is not expected to support high affinity binding of small molecules can nevertheless be targeted with potent inhibitors, and may be applicable to other targets conventionally considered undruggable. Citation Format: Rasmus Hansen, Ulf Peters, Anjali Babbar, Yuching Chen, Jun Feng, Matthew R. Janes, Liansheng Li, Pingda Ren, Yi Liu, Patrick P. Zarrinkar. Drugging an undruggable pocket: The biochemical mechanism of covalent KRASG12C inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 686.

Structural and solvent control of nonadiabatic photochemical bond formation: photocyclization of o-terphenyl in solution.

Elucidating the molecular dynamics that underlie photoinduced electrocyclization is a critical step toward controlling nonadiabatic photochemistry that enables bond formation. Here we present a comprehensive examination of the photochemical dynamics of o-terphenyl (OTP) in solution. Ultrafast transient absorption measurements demonstrate that OTP cyclizes upon 266 nm photoexcitation to form 4a,4b-dihydrotriphenylene (DHT) on a solvent-dependent time scale of 1.5-4 ps, considerably slower than the nonadiabatic cyclization of related diarylethenes. Correlations in these time scales versus bulk solvent properties reveal that mechanical rather than electrostatic solvent-solute interactions impact the excited-state relaxation rate, impeding nuclear dynamics leading toward the conical intersection for cyclization. In contrast, solvent-dependent mechanical interactions are observed to facilitate vibrational relaxation of DHT on time scales of 10-25 ps. DHT decays via thermally activated ring-opening with a lifetime of ∼46 ns in tetrahydrofuran, 12 orders of magnitude faster than dihydrophenanthrenes. We conclude that the differences in excited-state dynamics of OTP and diarylethenes and the relative stability of their cyclized products are determined by the relative strain induced by twisting the central carbon-carbon bond that bridges the terminal phenyl rings in each to enable bond formation. We relate these structure-dynamics relationships to the feasibility of photoinduced cyclodehydrogenation of o-arenes and design considerations for molecular photoswitches.

Adhesion Frequency Assay for <em>In Situ</em> Kinetics Analysis of Cross-Junctional Molecular Interactions at the Cell-Cell Interface

The micropipette adhesion assay was developed in 1998 to measure two-dimensional (2D) receptor-ligand binding kinetics. The assay uses a human red blood cell (RBC) as adhesion sensor and presenting cell for one of the interacting molecules. It employs micromanipulation to bring the RBC into contact with another cell that expresses the other interacting molecule with precisely controlled area and time to enable bond formation. The adhesion event is detected as RBC elongation upon pulling the two cells apart. By controlling the density of the ligands immobilized on the RBC surface, the probability of adhesion is kept in mid-range between 0 and 1. The adhesion probability is estimated from the frequency of adhesion events in a sequence of repeated contact cycles between the two cells for a given contact time. Varying the contact time generates a binding curve. Fitting a probabilistic model for receptor-ligand reaction kinetics to the binding curve returns the 2D affinity and off-rate. The assay has been validated using interactions of Fcγ receptors with IgG Fc, selectins with glycoconjugate ligands, integrins with ligands, homotypical cadherin binding, T cell receptor and coreceptor with peptide-major histocompatibility complexes. The method has been used to quantify regulations of 2D kinetics by biophysical factors, such as the membrane microtopology, membrane anchor, molecular orientation and length, carrier stiffness, curvature, and impingement force, as well as biochemical factors, such as modulators of the cytoskeleton and membrane microenvironment where the interacting molecules reside and the surface organization of these molecules. The method has also been used to study the concurrent binding of dual receptor-ligand species, and trimolecular interactions using a modified model. The major advantage of the method is that it allows study of receptors in their native membrane environment. The results could be very different from those obtained using purified receptors. It also allows study of the receptor-ligand interactions in a sub-second timescale with temporal resolution well beyond the typical biochemical methods. To illustrate the micropipette adhesion frequency method, we show kinetics measurement of intercellular adhesion molecule 1 (ICAM-1) functionalized on RBCs binding to integrin α(L)β(2) on neutrophils with dimeric E-selectin in the solution to activate α(L)β(2).