Ha-Jin Kim

Replication-Factor-C (RFC) and RFC-like complexes (RLCs) mediate chromatin engagement of the proliferating cell nuclear antigen (PCNA). It remains controversial how RFC and RLCs cooperate to regulate PCNA loading and unloading. Here, we show the distinct PCNA loading or unloading activity of each clamp loader. ATAD5-RLC possesses the potent PCNA unloading activity. ATPase motif and collar domain of ATAD5 are crucial for the unloading activity. DNA structures did not affect PCNA unloading activit

The physical properties of DNA have been suggested to play a central role in spatio-temporal organization of eukaryotic chromosomes. Experimental correlations have been established between the local nucleotide content of DNA and the frequency of inter- and intra-chromosomal contacts but the underlying physical mechanism remains unknown. Here, we combine fluorescence resonance energy transfer (FRET) measurements, precipitation assays, and molecular dynamics simulations to characterize the effect

Precision measurement is a hallmark of physics but the small length scale (∼nanometer) of elementary biological components and thermal fluctuations surrounding them challenge our ability to visualize their action. Here, we highlight the recent developments in single-molecule nanometry where the position of a single fluorescent molecule can be determined with nanometer precision, reaching the limit imposed by the shot noise, and the relative motion between two molecules can be determined with ∼0.

The current diagnosis of bacteremia mainly relies on blood culture, which is inadequate for the rapid and quantitative determination of most bacteria in blood. Here, a quantitative, multiplex, microfluidic fluorescence in situ hybridization method (μFISH) is developed, which enables early and rapid (3-h) diagnosis of bacteremia without the need for prior blood culture. This novel technology employs mannose-binding lectin-coated magnetic nanoparticles, which effectively opsonize a broad range of

Controlling efficiency and fidelity in the early stage of mitochondrial DNA transcription is crucial for regulating cellular energy metabolism. Conformational transitions of the transcription initiation complex must be central for such control, but how the conformational dynamics progress throughout transcription initiation remains unknown. Here, we use single-molecule fluorescence resonance energy transfer techniques to examine the conformational dynamics of the transcriptional system of yeast

Promoter recognition and local melting of DNA are key steps of transcription initiation catalyzed by RNA polymerase and initiation factors. From single molecule fluorescence resonance energy transfer studies of the yeast (Saccharomyces cerevisiae) mitochondrial RNA polymerase Rpo41 and its transcription factor Mtf1, we show that the pre-initiation complex is highly dynamic and undergoes repetitive opening-closing transitions that are modulated by Mtf1 and ATP. We found that Rpo41 alone has the i

The three-dimensional organization of chromatin and its time-dependent changes greatly affect virtually every cellular function, especially DNA replication, genome maintenance, transcription regulation, and cell differentiation. Sequencing-based techniques such as ChIP-seq, ATAC-seq, and Hi-C provide abundant information on how genomic elements are coupled with regulatory proteins and functionally organized into hierarchical domains through their interactions. However, visualizing the time-depen

Mitochondrial RNA polymerases depend on initiation factors, such as TFB2M in humans and Mtf1 in yeast Saccharomyces cerevisiae, for promoter-specific transcription. These factors drive the melting of promoter DNA, but how they support RNA priming and growth was not understood. We show that the flexible C-terminal tails of Mtf1 and TFB2M play a crucial role in RNA priming by aiding template strand alignment in the active site for high-affinity binding of the initiating nucleotides. Using single-m

The RecQ family of helicases is highly conserved both structurally and functionally from bacteria to humans. Defects in human RecQ helicases are associated with genetic diseases that are characterized by cancer predisposition and/or premature aging. RecQ proteins exhibit 3'-5' helicase activity and play critical roles in genome maintenance. Recent advances in single-molecule techniques have revealed the reiterative unwinding behavior of RecQ helicases. However, the molecular mechanisms involved

CRISPR-Cas-based genome imaging opened a new era of genome visualization in living cells. While genomic loci with repetitive sequences, such as centromeres and telomeres, can be reliably imaged, applying the technique to nonrepetitive genomic loci has remained challenging. Recent advancements in the design of CRISPR RNAs and Cas proteins, the development of novel fluorophores and the combination of CRISPR-Cas with other molecular machinery amplified target-specific signals and suppressed backgro

Abstract Controlling efficiency and fidelity in the early stage of mitochondrial DNA transcription is crucial for regulating cellular energy metabolism. Studies of bacteriophage and bacterial systems have revealed that transcription occurs through a series of conformational transitions during the initiation and elongation stages; however, how the conformational dynamics progress throughout these stages remains unknown. Here, we used single-molecule fluorescence resonance energy transfer techniqu

Longitudinal studies that analyze the changes in the axial diffusivity (AxD) and radial diffusivity (RD) values over time can elucidate the progression of white matter damage and its causal relationship with cognitive decline. This study aimed to investigate the longitudinal changes in white matter integrity based on AxD and RD and their association with cognitive decline in patients with mild cognitive impairment (MCI) that progressed to Alzheimer's disease (AD). Eighteen participants diagnosed

RecQ helicases are highly conserved between bacteria and humans. These helicases unwind various DNA structures in the 3' to 5'. Defective helicase activity elevates genomic instability and is associated with predisposition to cancer and/or premature aging. Recent single-molecule analyses have revealed the repetitive unwinding behavior of RecQ helicases from Escherichia coli to humans. However, the detailed mechanisms underlying this behavior are unclear. Here, we performed single-molecule studie