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Mobility Study of Individual Residue Sites in the Carbohydrate Recognition Domain of LSECtin Using SDSL–EPR Technique

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Abstract

Conformational changes in proteins profoundly influence their functional profiles. With site-directed spin labeling (SDSL)–electron paramagnetic resonance (EPR) spectroscopy, we investigated the mobility features of individual residue sites in the carbohydrate recognition domain (CRD) of LSECtin, a type II integral membrane protein. The mobility of six different residue sites scatting around the Ca2+-1-binding site were investigated by comparing their EPR spectra rotational correlation time τ c in order to obtain the information of conformational changes of relevant region. The results showed that the overall mobility of LSECtin-CRD increased after addition of Ca2+ and N-acetylglucosamine, but different sites in the CRD exhibited different mobility features, suggesting that these sites may have different functional profiles. The preliminary observations thus demonstrated that SDSL–EPR spectroscopy is not only an effective technique to reveal the mobility of single residue sites in LSECtin-CRD but also that the functions of single residue sites may be indicated by their conformational dynamics.

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Abbreviations

τ c :

Spectra rotational correlation time

CRD:

Carbohydrate recognition domain

DC:

Dendritic cell

EPR:

Electron paramagnetic resonance

GlcNAc:

N-Acetylglucosamine

IPTG:

Isopropyl-β-d-thiogalactoside

LSECtin:

Liver sinusoidal endothelial cell lectin

MTSL:

S-(2,2,5,5-Tetramethyl-2,5-dihydro-1H-pyrrol-3-yl) methyl methanesulfonothioate

SDSL:

Site-directed spin labeling

SOE-PCR:

Splicing by overlapped extension polymerase chain reaction

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Acknowledgments

This work is supported by the following funds: National Science Funds of China (no. 30970693 and no. 31170714).

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Authors and Affiliations

  1. Beijing Institute of Radiation Medicine, Beijing, China

    Changzhen Wang, Juntao Yang, Yu Zhou, Jianbo Cong, Guofu Dong, Xiangjun Hu, Li Tang & Ke Wu

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  1. Changzhen Wang
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  2. Juntao Yang
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  3. Yu Zhou
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  4. Jianbo Cong
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  5. Guofu Dong
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  6. Xiangjun Hu
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  7. Li Tang
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  8. Ke Wu
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Corresponding authors

Correspondence to Xiangjun Hu, Li Tang or Ke Wu.

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Changzhen Wang and Juntao Yang contributed equally to this work.

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Wang, C., Yang, J., Zhou, Y. et al. Mobility Study of Individual Residue Sites in the Carbohydrate Recognition Domain of LSECtin Using SDSL–EPR Technique. Appl Biochem Biotechnol 167, 2295–2304 (2012). https://doi.org/10.1007/s12010-012-9766-9

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