Viscoelasticity evolution in protein layers during binding reactions evaluated using high-frequency wireless and electrodeless quartz crystal microbalance biosensor without dissipation

In this study, we demonstrate the effectiveness of a resonance acoustic microbalance with a naked embedded quartz (RAMNE-Q) biosensor for evaluating viscoelastic property changes in thin protein layers during protein deposition reactions without dissipation measurement. Quartz crystal microbalance (QCM) biosensors have conventionally been adopted for the viscoelasticity evaluation of adsorbed protein layers by measuring dissipation as well as resonance frequency. However, dissipation, or the vibrational energy loss, is easily affected by many factors and is rarely measured with sufficiently high accuracy. To evaluate viscoelasticity only from a reliable frequency response, one needs to perform an ultrahighfrequency measurement, which is here achieved using the RAMNE-Q biosensor. Simultaneous frequency measurement is performed for fundamental and overtone modes up to 406MHz of a 58MHz RAMNE-Q biosensor during various binding reactions, and evolutions of viscosity, shear modulus, and thickness of adsorbed protein layers are inversely evaluated. A marked difference is observed in the viscosity evolution between specific and nonspecific binding reactions. Furthermore, the reversed frequency response appears, which indicates the modification of the protein structure into a rigid structure.Tomohiro Shagawa, Hiroomi Torii, Fumihito Kato, Hirotsugu Ogi and Masahiko Hirao. Viscoelasticity evolution in protein layers during binding reactions evaluated using high-frequency wireless and electrodeless quartz crystal microbalance biosensor without dissipation. Japanese Journal of Applied Physics, 2015, 54(9), 096601. https://doi.org/10.7567/JJAP.54.096601

Relationship between viscosity change and specificity in protein binding reaction studied by high-frequency wireless and electrodeless MEMS biosensor

This study proposes a methodology for evaluating specific binding behavior between proteins using a resonance acoustic microbalance with a naked-embedded quartz (RAMNE-Q) biosensor. We simultaneously measured the frequency responses of fundamental (58 MHz) and third-order (174 MHz) modes during multi step injections of proteins and deduced the thickness and viscosity evolutions of the protein layer. The viscosity increases with the progress of the binding reaction in nonspecific binding, but it markedly decreases in specific-binding cases. Thus, the high-frequency RAMNE-Q biosensor can be a powerful tool for evaluating specificity between proteins without measuring dissipation.Tomohiro Shagawa, Hiroomi Torii, Fumihito Kato, Hirotsugu Ogi and Masahiko Hirao. Relationship between viscosity change and specificity in protein binding reaction studied by high-frequency wireless and electrodeless MEMS biosensor. Japanese Journal of Applied Physics, 2015, 54(6), 068001. https://doi.org/10.7567/JJAP.54.068001

Data on the effect of hypomyelinating leukodystrophy 6 (HLD6)-associated mutations on the TUBB4A properties

Hypomyelinating leukodystrophy (HLD) is genetic demyelinating or dysmyelinating disease and is associated with at least 13 responsible genes. The mutations seem likely cause the functional deficiency of their gene products. HLD4- and HLD5-associated HSPD1 and FAM126A mutations affect biochemical properties of the gene products (Miyamoto et al. (2015,2014) [1,2]). Herein we provide the data regarding the effects of HLD6-associated tubulin beta 4A (TUBB4A) mutations on the properties

RNA Aptamer Binds Heparin-Binding Epidermal Growth Factor-Like Growth Factor with High Affinity and Specificity and Neutralizes Its Activity

Background: Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a member of the EGF family and is involved in various diseases including cancers. Aptamers are synthetic oligonucleotides (RNA or DNA) that fold into unique three-dimensional structures and specifically bind to their targets with high affinity. We aimed to generate an aptamer with high affinity and specificity for HB-EGF. Methods: Recombinant human HB-EGF (rhHB-EGF), comprised of the extracellular EGF-like and heparin-binding domains of HB-EGF, was used as the target. The aptamer against HB-EGF (the anti-HB-EGF aptamer) was obtained by systematic evolution of ligands by exponential enrichment (SELEX). Results: After the 10th round of SELEX, aptamers were reverse-transcribed and PCR-amplified. Within obtained forty-six clones, twenty-three were identical (the anti-HB-EGF aptamer). The analysis using wireless-electrodeless quartz crystal microbalance revealed that the anti-HB-EGF aptamer had high affinity for rhHB-EGF (KD value: 12.2 ± 1.1 nmol/L). The dot-blot analysis revealed that the anti-HB-EGF aptamer specifically bound to rhHB-EGF. The analysis using confocal microscopy indicated that the anti-HB-EGF aptamer also bound to membrane-bound HB-EGF. Western-blot assay indicated that the anti-HB-EGF aptamer inhibited the phosphorylation of rhHB-EGF-mediated EGF receptor (EGFR). Conclusion: We identified a novel RNA aptamer that bound with high affinity and specificity to rhHB-EGF and potently inhibited the rhHB-EGF-mediated phosphorylation of EGFR. The anti-HB-EGF aptamer may be a promising therapeutic agent for specifically neutralizing HB-EGF signaling