His-tag impact on structure.

Crystallographers are increasingly determining structures of protein constructs that include His tags. Many have taken for granted that these tags have little effect on the native structure. This paper surveys and compares crystal structures with and without His tags. It is observed that actual refined tag residues fitted into density occur in less that 10% of the tagged sequences. However, higher resolution crystals are observed when this occurs. It is shown that these purification tags generally have no significant effect on the structure of the native protein. Resolution and R factors are not affected, but the overall B factors are slightly higher. Additional annotation in the PDB format to make tag definition explicit is suggested.

[1]  H. Gilbert,et al.  How Family 26 Glycoside Hydrolases Orchestrate Catalysis on Different Polysaccharides , 2005, Journal of Biological Chemistry.

[2]  D. Waugh,et al.  Making the most of affinity tags. , 2005, Trends in biotechnology.

[3]  D. Hart,et al.  An approach to prevent aggregation during the purification and crystallization of wild type acyl coenzyme A: isopenicillin N acyltransferase from Penicillium chrysogenum. , 2005, Protein expression and purification.

[4]  Crystal structure of hypothetical protein PH0828 from Pyrococcus horikoshii , 2004, Proteins.

[5]  Zygmunt S Derewenda,et al.  The use of recombinant methods and molecular engineering in protein crystallization. , 2004, Methods.

[6]  R. Rudolph,et al.  Hexa‐histidin tag position influences disulfide structure but not binding behavior of in vitro folded N‐terminal domain of rat corticotropin‐releasing factor receptor type 2a , 2004, Protein science : a publication of the Protein Society.

[7]  I. Pikuleva,et al.  Expression of human cytochrome P450 46A1 in Escherichia coli: effects of N- and C-terminal modifications. , 2004, Archives of biochemistry and biophysics.

[8]  B. Kemper,et al.  N-terminal deletions and His-tag fusions dramatically affect expression of cytochrome p450 2C2 in bacteria. , 2001, Archives of biochemistry and biophysics.

[9]  T. N. Bhat,et al.  The Protein Data Bank , 2000, Nucleic Acids Res..

[10]  E. Myers,et al.  Basic local alignment search tool. , 1990, Journal of molecular biology.

[11]  C. Pidgeon,et al.  Chelating peptide-immobilized metal ion affinity chromatography. A new concept in affinity chromatography for recombinant proteins. , 1988, The Journal of biological chemistry.