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1.  Integrated solid-state nanopore platform for nanopore fabrication via dielectric breakdown, DNA-speed deceleration and noise reduction 
Scientific Reports  2016;6:31324.
The practical use of solid-state nanopores for DNA sequencing requires easy fabrication of the nanopores, reduction of the DNA movement speed and reduction of the ionic current noise. Here, we report an integrated nanopore platform with a nanobead structure that decelerates DNA movement and an insulating polyimide layer that reduces noise. To enable rapid nanopore fabrication, we introduced a controlled dielectric breakdown (CDB) process into our system. DNA translocation experiments revealed that single nanopores were created by the CDB process without sacrificing performance in reducing DNA movement speed by up to 10 μs/base or reducing noise up to 600 pArms at 1 MHz. Our platform provides the essential components for proceeding to the next step in the process of DNA sequencing.
PMCID: PMC4976334  PMID: 27499264
2.  Prevention of Dielectric Breakdown of Nanopore Membranes by Charge Neutralization 
Scientific Reports  2015;5:17819.
To achieve DNA sequencing using a solid-state nanopore, it is necessary to reduce the electric noise current. The noise current can be decreased by reducing the capacitance (C) of the nanopore device. However, we found that an electric-charge difference (ΔQ) between the electrolyte in one chamber and the electrolyte in another chamber occurred. For low capacitance devices, this electric-charge imbalance can lead to unexpectedly high voltage (ΔV = ΔQ/C) which disrupted the membrane when the two electrolytes were independently poured into the chambers. We elucidated the mechanism for the generation of initial defects and established new procedures for preventing the generation of defects by connecting an electric bypass between the chambers.
PMCID: PMC4669463  PMID: 26634995
3.  Exoproteome Profiles of Clostridium cellulovorans Grown on Various Carbon Sources 
Applied and Environmental Microbiology  2013;79(21):6576-6584.
The cellulosome is a complex of cellulosomal proteins bound to scaffolding proteins. This complex is considered the most efficient system for cellulose degradation. Clostridium cellulovorans, which is known to produce cellulosomes, changes the composition of its cellulosomes depending on the growth substrates. However, studies have investigated only cellulosomal proteins; profile changes in noncellulosomal proteins have rarely been examined. In this study, we performed a quantitative proteome analysis of the whole exoproteome of C. cellulovorans, including cellulosomal and noncellulosomal proteins, to illustrate how various substrates are efficiently degraded. C. cellulovorans was cultured with cellobiose, xylan, pectin, or phosphoric acid-swollen cellulose (PASC) as the sole carbon source. PASC was used as a cellulose substrate for more accurate quantitative analysis. Using an isobaric tag method and a liquid chromatography mass spectrometer equipped with a long monolithic silica capillary column, 639 proteins were identified and quantified in all 4 samples. Among these, 79 proteins were involved in saccharification, including 35 cellulosomal and 44 noncellulosomal proteins. We compared protein abundance by spectral count and found that cellulosomal proteins were more abundant than noncellulosomal proteins. Next, we focused on the fold change of the proteins depending on the growth substrates. Drastic changes were observed mainly among the noncellulosomal proteins. These results indicate that cellulosomal proteins were primarily produced to efficiently degrade any substrate and that noncellulosomal proteins were specifically produced to optimize the degradation of a particular substrate. This study highlights the importance of noncellulosomal proteins as well as cellulosomes for the efficient degradation of various substrates.
PMCID: PMC3811513  PMID: 23956399
4.  Profile of native cellulosomal proteins of Clostridium cellulovorans adapted to various carbon sources 
AMB Express  2012;2:37.
We performed a focused proteome analysis of cellulosomal proteins predicted by a genome analysis of Clostridium cellulovorans [Tamaru, Y., et al.. 2010. J. Bacteriol. 192:901–902]. Our system employed a long monolithic column (300 cm), which provides better performance and higher resolution than conventional systems. Twenty-three cellulosomal proteins were, without purification, identified by direct analysis of the culture medium. Proteome analysis of the C. cellulovorans cellulosome after culture in various carbon sources demonstrated the production of carbon source-adapted cellulosome components.
PMCID: PMC3444338  PMID: 22839966
Clostridium cellulovorans; Cellulosome; Focused proteome analysis; Monolithic column

Results 1-4 (4)