当前位置： SCI文献检索 > BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS期刊下所有文献 > Mutation in the β-hairpin of the Bordetella pertussis adenylate cyclase toxin modulates N-lobe conformation in calmodulin.

Mutation in the β-hairpin of the Bordetella pertussis adenylate cyclase toxin modulates N-lobe conformation in calmodulin.

Abstract：

:Bordetella pertussis, causative agent of whooping cough, produces an adenylate cyclase toxin (CyaA) that is an important virulence factor. In the host cell, the adenylate cyclase domain of CyaA (CyaA-ACD) is activated upon association with calmodulin (CaM), an EF-hand protein comprised of N- and C-lobes (N-CaM and C-CaM, respectively) connected by a flexible tether. Maximal CyaA-ACD activation is achieved through its binding to both lobes of intact CaM, but the structural mechanisms remain unclear. No high-resolution structure of the intact CaM/CyaA-ACD complex is available, but crystal structures of isolated C-CaM bound to CyaA-ACD shed light on the molecular mechanism by which this lobe activates the toxin. Previous studies using molecular modeling, biochemical, and biophysical experiments demonstrate that CyaA-ACD's β-hairpin participates in site-specific interactions with N-CaM. In this study, we utilize nuclear magnetic resonance (NMR) spectroscopy to probe the molecular association between intact CaM and CyaA-ACD. Our results indicate binding of CyaA-ACD to CaM induces large conformational perturbations mapping to C-CaM, while substantially smaller structural changes are localized primarily to helices I, II, and IV, and the metal-binding sites in N-CaM. Site-specific mutations in CyaA-ACD's β-hairpin structurally modulate N-CaM, resulting in conformational perturbations in metal binding sites I and II, while no significant structural modifications are observed in C-CaM. Moreover, dynamic light scattering (DLS) analysis reveals that mutation of the β-hairpin results in a decreased hydrodynamic radius (Rh) and reduced thermal stability in the mutant complex. Taken together, our data provide new structural insights into the β-hairpin's role in stabilizing interactions between CyaA-ACD and N-CaM.

journal_name

Biochem Biophys Res Commun

journal_title

Biochemical and biophysical research communications

authors

Springer TI,Goebel E,Hariraju D,Finley NL

doi

10.1016/j.bbrc.2014.09.048

subject

Has Abstract

pub_date

2014-10-10 00:00:00

pages

43-8

issue

eissn

0006-291X

issn

1090-2104

pii

S0006-291X(14)01664-7

journal_volume

453

pub_type

杂志文章

在线工具

Mutation in the β-hairpin of the Bordetella pertussis adenylate cyclase toxin modulates N-lobe conformation in calmodulin.