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Expression and Purification of human Neuronal PAS domain protein 2 (hNPAS2)
Author(s) -
Moriel Karla,
Xi Dongman,
Sarabia Alejandro,
Chauhan Neha,
Ray Supriyo,
Xiao Chuan
Publication year - 2018
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2018.32.1_supplement.526.15
Subject(s) - circadian rhythm , circadian clock , biology , microbiology and biotechnology , transcription (linguistics) , regulator , transcription factor , homogeneous , gene , computational biology , neuroscience , genetics , philosophy , linguistics , physics , thermodynamics
The circadian rhythm is an internal clock responsible for a variety of physiological processes occurring within a 24‐hour cycle. Synchronized by a master clock in the brain, the circadian rhythm consists of a transcriptional‐translational feedback loop (TTFL) capable of inducing oscillating gene expression in all the cells throughout the organism. The TTFL begins with the heterodimerization of two transcription factors, BMAL1 and CLOCK, resulting in the transcription and translation of many circadian‐related genes. The target protein of interest, human neuronal PAS domain protein 2 (hNPAS2) is a paralog of CLOCK that can serve as a positive regulator of the circadian rhythm by heterodimerizing with BMAL1. Previous research suggests that disruption in the expression of hNPAS2 may result in improper circadian functioning and development of sleeping and mental disorders. In this study, the full‐length eukaryotic form of hNPAS2 was cloned into a bacterial expression vector. However, bacterial expression has led to misfolding and aggregation of the recombinant hNPAS2, becoming the central issue of the study. In an effort to obtain soluble and homogeneous hNPAS2, multiple expression and purification methods have been developed. The resultant purified soluble protein will enable future structural and functional studies, which can bring new insights to the regulatory mechanisms of the circadian rhythm and understanding of circadian‐related disorders. Additionally, a portion of this research was integrated into a teaching module delivered by the leading student researcher to a senior undergraduate advanced biochemistry course. Multiple novel pedagogic techniques were employed in the teaching module with the objective of raising the students' interests to research as well as their abilities to apply the principles and concepts of biochemistry learned in the course in a real scientific study. Support or Funding Information Research reported in this poster was supported by the National Institute of General medical Sciences of the National Institutes of Health under linked Award Numbers RL5GM118969, TL4GM118971, AND UL1GM118970. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health Research reported in this poster was supported by the National Research of General Medical Sciences of NIH under award number SC3GM109870 Research reported in this poster was supported has used core facility at BBRC supported by National Institute of Minority Health and Health Disparities under award number 5G12MD007592 This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .