Nickel Binding and [NiFe]-Hydrogenase Maturation by the Metallochaperone SlyD with a Single Metal-Binding Site in Escherichia coli

• Published in: Journal of molecular biology 2012-03, Vol.417 (1-2), p.28-35
• Main Authors: Kaluarachchi, Harini, Altenstein, Matthias, Sugumar, Sonia R., Balbach, Jochen, Zamble, Deborah B., Haupt, Caroline
• Format: Article
• Language: English
• Subjects: absorption; Amino Acid Motifs; Binding Sites; biogenesis; biosynthesis; calorimetry; conserved single nickel-binding motif; crosslinking; electrospray ionization mass spectrometry; Escherichia coli; Escherichia coli - chemistry; Escherichia coli - enzymology; Escherichia coli Proteins - chemistry; Escherichia coli Proteins - metabolism; ferredoxin hydrogenase; histidine; Hydrogenase - chemistry; Hydrogenase - metabolism; HypB; ions; Ligands; metal-binding domain; Metallochaperones - chemistry; Metallochaperones - metabolism; Metals - chemistry; Metals - metabolism; nickel; NMR spectroscopy; nuclear magnetic resonance spectroscopy; Peptidylprolyl Isomerase - chemistry; Peptidylprolyl Isomerase - metabolism; Spectrometry, Mass, Electrospray Ionization; Thermus thermophilus; titration; X-radiation; X-ray absorption spectroscopy
• ISSN: 0022-2836; 1089-8638
• DOI: 10.1016/j.jmb.2012.01.037

SlyD (sensitive to lysis D) is a nickel metallochaperone involved in the maturation of [NiFe]-hydrogenases in Escherichia coli (E. coli) and specifically contributes to the nickel delivery step during enzyme biosynthesis. This protein contains a C-terminal metal-binding domain that is rich in potential metal-binding residues that enable SlyD to bind multiple nickel ions with high affinity. The SlyD homolog from Thermus thermophilus does not contain the extended cysteine- and histidine-rich C-terminal tail of the E. coli protein, yet it binds a single Ni(II) ion tightly. To investigate whether a single metal-binding motif can functionally replace the full-length domain, we generated a truncation of E. coli SlyD, SlyD155. Ni(II) binding to SlyD155 was investigated by using isothermal titration calorimetry, NMR and electrospray ionization mass spectrometry measurements. This in vitro characterization revealed that SlyD155 contains a single metal-binding motif with high affinity for nickel. Structural characterization by X-ray absorption spectroscopy and NMR indicated that nickel was coordinated in an octahedral geometry with at least two histidines as ligands. Heterodimerization between SlyD and another hydrogenase accessory protein, HypB, is essential for optimal hydrogenase maturation and was confirmed for SlyD155 via cross-linking experiments and NMR titrations, as were conserved chaperone and peptidyl-prolyl isomerase activities. Although these properties of SlyD are preserved in the truncated version, it does not modulate nickel binding to HypB in vitro or contribute to the maturation of [NiFe]-hydrogenases in vivo, unlike the full-length protein. This study highlights the importance of the unusual metal-binding domain of E. coli SlyD in hydrogenase biogenesis. [Display omitted] ► Ni(II) is tightly bound by the histidines of the metal-binding motif. ► Complex formation between SlyD155 [IF (insert in flap) domain] and HypB was confirmed in vitro. ► SlyD155 is not as effective as full-length SlyD in inducing Ni(II) release from HypB. ► SlyD155 cannot stimulate hydrogenase activity in vivo. ► The entire metal-binding domain holds a major role in Ni(II) sequestration.