A novel thermostable hemoglobin from the actinobacterium Thermobifida fusca.
The gene coding for a hemoglobin-like protein (Tf-trHb) has been identified in the thermophilic actinobacterium Thermobifida fusca and cloned in Escherichia coli for overexpression. The crystal structure of the ferric, acetate-bound derivative, was obtained at 2.48 A resolution. The three-dimensional structure of Tf-trHb is similar to structures reported for the truncated hemoglobins from Mycobacterium tuberculosis and Bacillus subtilis in its central domain. The complete lack of diffraction patterns relative to the N- and C-terminal segments indicates that these are unstructured polypeptides chains, consistent with their facile cleavage in solution. The absence of internal cavities and the presence of two water molecules between the bound acetate ion and the protein surface suggest that the mode of ligand entry is similar to that of typical hemoglobins. The protein is characterized by higher thermostability than the similar mesophilic truncated hemoglobin from B. subtilis, as demonstrated by far-UV CD melting experiments on the cyano-met derivatives. The ligand-binding properties of Tf-trHb, analyzed in stopped flow experiments, demonstrate that Tf-trHb is capable of efficient O2 binding and release between 55 and 60 degrees C, the optimal growth temperature for Thermobifida fusca.
Bonamore
Alessandra
A
Department of Biochemical Sciences, University of Rome La Sapienza, Italy.
Ilari
Andrea
A
Giangiacomo
Laura
L
Bellelli
Andrea
A
Morea
Veronica
V
Boffi
Alberto
A
D039903:Actinobacteria / Q000737:chemistry*; D000595:Amino Acid Sequence; D002942:Circular Dichroism; D003001:Cloning, Molecular; D006454:Hemoglobins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D008024:Ligands; D008969:Molecular Sequence Data; D010100:Oxygen / Q000378:metabolism; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D013056:Spectrophotometry, Ultraviolet; D013696:Temperature; D054793:Truncated Hemoglobins
D006454:Hemoglobins; D008024:Ligands; D054793:Truncated Hemoglobins; D010100:Oxygen