Proteins often require cofactors to perform their biological functions and must fold in the presence of their cognate ligands. Using circular dichroism spectroscopy. we investigated the effects of divalent metal binding upon the folding pathway of Escherichia coli RNase HI. This enzyme binds divalent metal in its active site, which is proximal to the folding core of RNase HI as defined by hydro...

The LAGLIDADG homing endonucleases include free-standing homodimers, pseudosymmetric monomers, and related enzyme domains embedded within inteins. DNA-bound structures of homodimeric I-CreI and monomeric I-SceI indicate that three catalytic divalent metal ions are distributed across a pair of overlapping active sites, with one shared metal participating in both strand cleavage reactions. These ...

The SmtB/ArsR family of prokaryotic metalloregulatory transcriptional repressors represses the expression of operons linked to stress-inducing concentrations of di- and multivalent heavy metal ions. Derepression results from direct binding of metal ions by these homodimeric "metal sensor" proteins. An evolutionary analysis, coupled with comparative structural and spectroscopic studies of six Sm...

Lmb is a 34 kDa laminin binding surface adhesin of Streptococcus agalactiae. The structure of Lmb reported by us recently has shown that it consists of a metal binding crevice, in which a zinc ion is coordinated to three highly conserved histidines. To elucidate the structural and functional significance of the metal ion in Lmb, these histidines have been mutated to alanine and single, double a...

Binding of a transition metal ion to specific sites in concanavalin A induces the formation of specific Ca(2+) ion-binding sites. Sites for binding alpha-methyl d-glucopyranoside exist only when a transition metal ion and Ca(2+) ion are bound.

To understand the key determinants in calcium-binding affinity, a calcium-binding site with pentagonal bipyramid geometry was designed into a non-calcium-binding protein, domain 1 of CD2. This metal-binding protein has five mutations with a net charge in the coordination sphere of -5 and is termed DEEEE. Fluorescence resonance energy transfer was used to determine the metal-binding affinity of ...

Using a cloned single domain of the high mobility group protein 1 (HMGB1), we evaluated the effect of introducing metal binding site(s) on protein stability and function. An HMG domain is a conserved sequence of approximately 80 amino acids rich in basic, aromatic and proline residues that is active in binding DNA in a sequence- or structure-specific manner. The design strategy focuses on ancho...

NAD-dependent isocitrate dehydrogenase from pig heart contains three types of subunits of slightly different molecular weights in an approximate ratio of 2:l:l. Ultrafiltration binding experiments at pH 6.1 indicate 1 binding site for every 2 subunits for several different ligands. The metal activator, Mn2+, binds to isocitrate dehydrogenase in the absence of other ligands ( K D = 115 p ~ ) . M...

The adhesiveness of integrin L 2 is modulated by divalent cations. We mutated three metal ion-binding sites in the 2 I domain. The metal ion-dependent adhesion site (MIDAS) and the ligandinduced metal-binding site are required for ligand binding and sufficient for synergism between Ca2 and Mg2 . Adjacent to MIDAS (ADMIDAS) mutants are constitutively active but remain bent, with poor exposure of...

BACKGROUND The pathological hallmark of Parkinson's disease is the deposition of cytoplasmic neuronal inclusions termed Lewy bodies. The major component of Lewy bodies is amyloid fibrils of α-synuclein. To investigate what causes α-synuclein aggregation is essential to understand its pathological roles in Parkinson's disease. Various metal ions, including iron and copper, have been implicated i...