Independent Movement, Dimerization and Stability of Tandem Repeats of Chicken Brain α-Spectrin

Kusunoki, Hideki; Minasov, G.; MacDonald, Robert C.; Mondragón, Alfonso

doi:10.1016/j.jmb.2004.09.019

Cited by 96 publications

(115 citation statements)

References 70 publications

(95 reference statements)

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“…The calculated spectra were indistinguishable from those measured (not shown). The discrepancy between the modeled sum of helical conformations equaling 94% and the helical regions in X-ray crystal structures of repeats equaling 87% (25,33,34) may reflect the assignment of some loop regions as distorted helix in the latter.…”

Section: Both Zu5 and 272 And Their Parent Spectrin Binding Domain Ofmentioning

confidence: 99%

“…This reasoning also applies, of course, to the other prominent members of the spectrin family, α-actinin and dystrophin (30). It remains a mystery, however, as to what dictates this structural versatility of the signature spectrin repeating unit motif (31) in spite of the availability of Xray crystal structures of two (25,32,33) and three (34) repeat fragments of spectrin and α-actinin. An exception is the structure of four repeat fragments of α-actinin (35) with a lengthwise twist which enables interaction of α-actinin with actin at each end.…”

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confidence: 99%

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Molecular Epitopes of the Ankyrin−Spectrin Interaction

et al. 2008

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Isoforms of ankyrin and its binding partner spectrin are responsible for a number of interactions in a variety of human cells. Conflicting evidence, however, had identified two different, non-overlapping human erythroid ankyrin subdomains, Zu5 and 272, as the minimum binding region for β-spectrin. Complementary studies on the ankyrin-binding domain of spectrin have been somewhat more conclusive yet have not presented binding in terms of well-phased, integral numbers of spectrin repeats. Thus, the objective of this study was to clearly define and characterize the minimal ankyrin−spectrin binding epitopes. Circular dichroism (CD) wavelength spectra of the aforementioned ankyrin subdomains show that these fragments are 30−60% unstructured. In contrast, human erythroid β-spectrin repeats 13, 14, 15, and 16 (prepared in all combinations of two adjacent repeats) demonstrated proper folding and stability as determined by CD and tryptophan wavelength and heat denaturation scans. Native polyacrylamide gel electrophoresis (PAGE) gel shifts as well as affinity pull-down assays implicated Zu5 and β-spectrin repeats 14−15 as the minimum binding epitopes. These results were confirmed by analytical ultracentrifugation to sedimentation equilibrium by which a 1:1 complex was obtained if and only if Zu5 was mixed with β-spectrin constructs containing repeats 14 and 15 in tandem. Surface plasmon resonance yielded a K D of 15.2 nM for binding of β-spectrin fragments to the ankyrin subdomain Zu5, accounting for all of the binding observed between the intact molecules. Collectively, these results show the 14th and 15th β-spectrin repeats comprise the minimal, phased region of β-spectrin, which binds ankyrin at the Zu5 subdomain with high affinity.

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Section: Both Zu5 and 272 And Their Parent Spectrin Binding Domain Ofmentioning

confidence: 99%

mentioning

confidence: 99%

Molecular Epitopes of the Ankyrin−Spectrin Interaction

et al. 2008

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“…free energy landscape | frustration | phi value | protein folding T he 15th, 16th, and 17th domains of chicken brain α-spectrin (R15, R16, and R17) have very similar structures, stabilities, and β-Tanford (β T ) values (which reflect the compactness of the transition state for folding and unfolding) (1)(2)(3)(4)(5)(6). However, the folding of R15 differs from that of R16 and R17 in a number of respects.…”

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confidence: 99%

Separating the effects of internal friction and transition state energy to explain the slow, frustrated folding of spectrin domains

Wensley

Kwa

Shammas

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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The elongated three-helix bundle domains spectrin R16 and R17 fold some two to three orders of magnitude more slowly than their homologue R15. We have shown that this slow folding is due, at least in part, to roughness in the free-energy landscape of R16 and R17. We have proposed that this roughness is due to a frustrated search for the correct docking of partly preformed helices. However, this accounts for only a small part of the slowing of folding and unfolding. Five residues on the A helix of R15, when inserted together into R16 or R17, increase the folding rate constants, reduce landscape roughness, and alter the folding mechanism to one resembling R15. The effect of each of these mutations individually is investigated here. No one mutation causes the behavior seen for the five in combination. However, two mutations, E18F and K25V, significantly increase the folding and unfolding rates of both R16 and R17 but without a concomitant loss in landscape roughness. E18F has the greatest effect on the kinetics, and a Φ-value analysis of the C helix reveals that the folding mechanism is unchanged. For both E18F and K25V the removal of the charge and resultant transition state stabilization is the main origin of the faster folding. Consequently, the major cause of the unusually slow folding of R16 and R17 is the non-native burial of the two charged residues in the transition state. The slowing due to landscape roughness is only about fivefold.free energy landscape | frustration | phi value | protein folding T he 15th, 16th, and 17th domains of chicken brain α-spectrin (R15, R16, and R17) have very similar structures, stabilities, and β-Tanford (β T ) values (which reflect the compactness of the transition state for folding and unfolding) (1-6). However, the folding of R15 differs from that of R16 and R17 in a number of respects. R15 folds and unfolds two orders of magnitude faster than R16 and three orders of magnitude faster than R17. R16 and R17 have two sequential transition states, and for both domains the first of these (TS1) shows significant landscape roughness (or "internal friction") (7). This has not been seen for any other domain of comparable size and folding kinetics, although theory has long predicted the possibility of such landscape roughness (8-15). R15 has a broad transition state (characterized by "rollover" in the unfolding limb for some mutants and for wild type under some conditions), but due to the speed of folding and unfolding it is not known whether it has two sequential transition states (16). However, the early transition state of R15 (which corresponds to TS1 of R16 and R17 and will be referred to as such) has a smoother, less frustrated landscape (7).Φ-value analysis shows that for all three domains the A and C helices are partially structured at TS1, whereas the B helix is relatively unstructured (16-18). R16 and R17 fold via a framework-like mechanism, with some tertiary contacts formed but more extensive secondary structure that extends throughout the A and C helices. In contrast, R1...

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“…These characteristics also make spectrin flexible [13]. The X-ray crystallographic data for the chicken-brain -spectrin triple -helix repeat bundle indicate that the connections between spectrin repeats can bend independently without affecting the adjacent connection [14]. Recently, our group determined the medium resolution three-dimensional structures of a spectrin dimer initiation site and a tetrameric head-head interaction site, using homology modeling and chemical cross-linking combined with mass spectrometry [1518].…”

Section: The Structure Of Spectrinmentioning

confidence: 99%

Spectrin: Structure, function and disease

Zhang

Zhao

et al. 2013

Sci. China Life Sci.

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Spectrin is a large, cytoskeletal, and heterodimeric protein composed of modular structure of and subunits, it typically contains 106 contiguous amino acid sequence motifs called "spectrin repeats". Spectrin is crucial for maintaining the stability and structure of the cell membrane and the shape of a cell. Moreover, it contributes to diverse cell functions such as cell adhesion, cell spreading, and the cell cycle. Mutations of spectrin lead to various human diseases such as hereditary hemolytic anemia, type 5 spinocerebellar ataxia, cancer, as well as others. This review focuses on recent advances in determining the structure and function of spectrin as well as its role in disease.erythrocyte, spectrin, cell cycle, mass spectrometry, disease Citation:Zhang R, Zhang C Y, Zhao Q, et al. Spectrin: Structure, function and disease.

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Independent Movement, Dimerization and Stability of Tandem Repeats of Chicken Brain α-Spectrin

Cited by 96 publications

References 70 publications

Molecular Epitopes of the Ankyrin−Spectrin Interaction

Molecular Epitopes of the Ankyrin−Spectrin Interaction

Separating the effects of internal friction and transition state energy to explain the slow, frustrated folding of spectrin domains

Spectrin: Structure, function and disease

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