NMR and crystallographic structural studies of the extremely stable monomeric variant of human cystatin C with single amino acid substitution

Martyna Maszota-Zieleniak, Przemyslaw Jurczak, Marta Orlikowska, Igor Zhukov, Dominika Borek, Zbyszek Otwinowski, Piotr Skowron, Zuzanna Pietralik, Maciej Kozak, Aneta Szymańska, Sylwia Rodziewicz-Motowidło

Research output: Contribution to journalArticle

1 Scopus citations

Abstract

Human cystatin C (hCC), a member of the superfamily of papain-like cysteine protease inhibitors, is the most widespread cystatin in human body fluids. This small protein, in addition to its physiological function, is involved in various diseases, including cerebral amyloid angiopathy, cerebral hemorrhage, stroke, and dementia. Physiologically active hCC is a monomer. However, all structural studies based on crystallization led to the dimeric structure formed as a result of a three-dimensional exchange of the protein domains (3D domain swapping). The monomeric structure was obtained only for hCC variant V57N and for the protein stabilized by an additional disulfide bridge. With this study, we extend the number of models of monomeric hCC by an additional hCC variant with a single amino acid substitution in the flexible loop L1. The V57G variant was chosen for the X-ray and NMR structural analysis due to its exceptional conformational stability in solution. In this work, we show for the first time the structural and dynamics studies of human cystatin C variant in solution. We were also able to compare these data with the crystal structure of the hCC V57G and with other cystatins. The overall cystatin fold is retained in the solute form. Additionally, structural information concerning the N terminus was obtained during our studies and presented for the first time. Database: Crystallographic structure: structural data are available in PDB databases under the accession number 6ROA. NMR structure: structural data are available in PDB and BMRB databases under the accession numbers 6RPV and 34399, respectively.

Original languageEnglish (US)
Pages (from-to)361-376
Number of pages16
JournalFEBS Journal
Volume287
Issue number2
DOIs
StatePublished - Jan 1 2020

Keywords

  • NMR
  • X-ray
  • hCC V57G variant
  • human cystatin C
  • protein structure

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Maszota-Zieleniak, M., Jurczak, P., Orlikowska, M., Zhukov, I., Borek, D., Otwinowski, Z., Skowron, P., Pietralik, Z., Kozak, M., Szymańska, A., & Rodziewicz-Motowidło, S. (2020). NMR and crystallographic structural studies of the extremely stable monomeric variant of human cystatin C with single amino acid substitution. FEBS Journal, 287(2), 361-376. https://doi.org/10.1111/febs.15010