Local structural preferences of calpastatin, the intrinsically unstructured protein inhibitor of calpain

Robert Kiss, Dénes Kovács, Péter Tompa, A. Perczel

Research output: Contribution to journalArticle

39 Citations (Scopus)

Abstract

Calpain, the calcium-activated intracellular cysteine protease, is under the tight control of its intrinsically unstructured inhibitor, calpastatin. Understanding how potent inhibition by calpastatin can be reconciled with its unstructured nature provides deeper insight into calpain function and a more general understanding of how proteins devoid of a well-defined structure carry out their function. To this end, we performed a full NMR assignment of hCSD1 to characterize it in its solution state. Secondary chemical shift values and NMR relaxation data, R1, R2, and hetero-NOE, as well as spectral density function analysis have shown that conserved regions of calpastatin, subdomains A and C, which are responsible for calcium-dependent anchoring of the inhibitor to the enzyme, preferentially sample partially helical backbone conformations of a reduced flexibility. Moreover, the linker regions between subdomains are more flexible with no structural preference. The primary determinant of calpain inhibition, subdomain B, also has a non-fully random conformational preference, resembling a β-turn structure also ascertained by prior studies of a 27-residue peptide encompassing the inhibitory region. This local structural preference is also confirmed by a deviation in chemical shift values between full-length calpastatin domain 1 and a truncated construct cut in the middle of subdomain B. At the C-terminal end of the molecule, a nascent helical region was found, which in contrast to the overall structural properties of the molecule may indicate a previously unknown functional region. Overall, these observations provide further evidence that supports previous suggestions that intrinsically unstructured proteins use preformed structural elements in efficient partner recognition.

Original languageEnglish
Pages (from-to)6936-6945
Number of pages10
JournalBiochemistry
Volume47
Issue number26
DOIs
Publication statusPublished - Jul 1 2008

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Intrinsically Disordered Proteins
Calpain
Chemical shift
Nuclear magnetic resonance
Calcium
Molecules
Cysteine Proteases
Spectral density
Enzyme Inhibitors
Probability density function
Conformations
Structural properties
Peptides
calpain inhibitors
calpastatin
Enzymes
Proteins

ASJC Scopus subject areas

  • Biochemistry

Cite this

Local structural preferences of calpastatin, the intrinsically unstructured protein inhibitor of calpain. / Kiss, Robert; Kovács, Dénes; Tompa, Péter; Perczel, A.

In: Biochemistry, Vol. 47, No. 26, 01.07.2008, p. 6936-6945.

Research output: Contribution to journalArticle

Kiss, Robert ; Kovács, Dénes ; Tompa, Péter ; Perczel, A. / Local structural preferences of calpastatin, the intrinsically unstructured protein inhibitor of calpain. In: Biochemistry. 2008 ; Vol. 47, No. 26. pp. 6936-6945.
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