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Identification of critical amino acid residues and functional conservation of the Neurospora crassa and Rattus norvegicus orthologues of neuronal calcium sensor-1

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Abstract

Neuronal calcium sensor-1 (NCS-1) is a member of neuronal calcium sensor family of proteins consisting of an amino terminal myristoylation domain and four conserved calcium (Ca2+) binding EF-hand domains. We performed site-directed mutational analysis of three key amino acid residues that are glycine in the conserved site for the N-terminal myristoylation, a conserved glutamic acid residue responsible for Ca2+ binding in the third EF-hand (EF3), and an unusual non-conserved amino acid arginine at position 175 in the Neurospora crassa NCS-1. The N. crassa strains possessing the ncs-1 mutant allele of these three amino acid residues showed impairment in functions ranging from growth, Ca2+ stress tolerance, and ultraviolet survival. In addition, heterologous expression of the NCS-1 from Rattus norvegicus in N. crassa confirmed its interspecies functional conservation. Moreover, functions of glutamic acid at position 120, the first Ca2+ binding residue among all the EF-hands of the R. norvegicus NCS-1 was found conserved. Thus, we identified three critical amino acid residues of N. crassa NCS-1, and demonstrated its functional conservation across species using the orthologue from R. norvegicus.

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Acknowledgments

DG and RD were supported by Research Fellowships, respectively, from the Ministry of Human Resource Development (MHRD) and Council of Scientific and Industrial Research-University Grant Commission (CSIR-UGC), Government of India. We thank Prof. R. D. Burgoyne (University of Liverpool, UK) for kindly providing us the pET5α- ncs-1 Rat construct containing the R. norvegicus ncs-1 cDNA. This work was supported partially by a Grant (BT/PR3635/BCE/8/892/2012) to RT from the Department of Biotechnology, Government of India. The FGSC generously waived charges for strains and race tubes. The FGSC was supported by NSF Grant BIR-9222772.

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    1. Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati, 781 039, India

      Dibakar Gohain, Rekha Deka & Ranjan Tamuli

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    1. Dibakar Gohain
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    2. Rekha Deka
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    Correspondence to Ranjan Tamuli.

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    Dibakar Gohain and Rekha Deka have contributed equally to this work.

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    Gohain, D., Deka, R. & Tamuli, R. Identification of critical amino acid residues and functional conservation of the Neurospora crassa and Rattus norvegicus orthologues of neuronal calcium sensor-1. Genetica 144, 665–674 (2016). https://doi.org/10.1007/s10709-016-9933-y

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