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A de novo designed calcium-binding site into the framework of a non-calcium-binding protein, domain 1 of CD2, is introduced. The feasibility of designing a single calcium-binding site into a host protein, taking into account only local properties of a calcium-binding site obtained by a survey of natural calcium-binding proteins and chelators has been demonstrated.

The rational design of a novel class of magnetic resonance imaging (MRI) contrast agents with engineered proteins chelated with gadolinium is described. The agents have displayed a 20-fold increase in longitudinal and transverse relaxation rate values and the protein contrast agents are used as molecular imaging probes to target disease markers, which have extended applications of MRI.

Calcium ions play key roles as structural components in biomineralization and as a second messenger in signaling pathways. We have introduced a de novo designed calcium-binding site into the framework of a non-calcium-binding protein, domain 1 of CD2. The resulting protein selectively binds calcium over magnesium with calcium-binding affinity comparable to that of natural extracellular calcium-binding proteins (K[sub d] of 50 µM). This experiment is the first successful metalloprotein design that has a high coordination number (seven) metal-binding site constructed into a β-sheet protein. Our results demonstrate the feasibility of designing a single calcium-binding site into a host protein, taking into account only local properties of a calcium-binding site obtained by a survey of natural calcium-binding proteins and chelators. The resulting site exhibits strong metal selectivity, suggesting that it should now be feasible to understand and manipulate signaling processes by designing novel calcium-modulated proteins with specifically desired functions and to affect their stability. [ABSTRACT FROM AUTHOR]