FusionTagsUsedforRecombinantProteinExpressionand.docVIP

Fusion Tags Used for Recombinant Protein Expression and Purification* Tag Size Fusion tag location Tag type Comments/Remarks His-tag 6, 8, or 10 aa N-, C-, internal Purification Most common purification tag used for immobilized metal affinity chromatography (IMAC) one-step purification [81]. Purification possible even under denaturing conditions [82]. Tag possibly influences crystallization T7-tag 11 or 16 aa N-, internal Purification, enhanced expression Monoclonal antibody-based purification (denaturing low pH elution needed). Leaves unnatural N-terminal amino acids on the recombinant protein. Possibly enhanced expression levels since the T7-tag is derived from the T7 gene 10 which is the naturally most abundant phage T7 gene product. S-tag 15 aa N-, C-, internal Purification and detection S-protein (104 aa, Ribonuclease A minus S-tag peptide sequence) modified resin affinity purification. RNAse S assay possible for quantitative assay of expression levels. FLAG( peptide (DYKDDDDK) 8 aa N-,C- Purification Ca2+-dependent monoclonal antibody affinity purification with EDTA elution. Tag cleavable with enterokinase [83]. thioredoxin 109 aa (11.7 kDa) N-,C- Purification and enhanced expression Affinity purification with phenylarsine oxide-modified (ThioBond) resin. His-patch thioredoxin 109 aa (11.7 kDa) N-,C- Purification and enhanced expression Use of His-patch modified thioredoxin for IMAC affinity purification [84]. lacZ ((-Galactosidase) 116 kDa N-,C- Purification Purification using p-amino-phenyl-(-D-thiogalactoside-modified sepharose. Classical tag used for protecting peptides from proteolytic degradation. However, fusion proteins with this tag have a high tendency to be insoluble. Active enzyme is a tetramer chloramphenicol acetyltransferase 24 kDa N- Secretion, purification and detection Chloramphenicol-sepharose purification. Enzymatic assay possible for quantitation. trpE 27 kDa N- Purification Often form insoluble precipitates. Hydrophobic interac