Twigged streptavidin polymer as a scaffold for protein assembly

Broadening the scope of sortagging

“…This novel approach for protein engineering was extensively expanded in the following years, e.g. for the ligation of uorescent proteins to a protein of interest, [36][37][38] to develop immunoassays with the aim of monitoring glucose levels, 39 to crosslink enzymes, 40 to generate fusion proteins that are inaccessible via direct expression, 41 to form an enzyme cascade within E. coli 42 and to ligate two domains of a protein. 43,44 Other studies focused on the ligation of two peptides [45][46][47] and of a peptide to a protein, [48][49][50][51][52][53] among others to demonstrate the purication of recombinant fusion proteins, 54 to functionalize amyloid-based polymers, 55 to assemble semisynthetic proteins or protein mimics 56,57 and to incorporate functional proteins into hydrogels.…”

Natural Occurring and Engineered Enzymes for Peptide Ligation and Cyclization

“…Already in 2004 sortagging was described for protein/peptide ligation to another protein/peptide, even containing unnatural AAs (Mao et al, 2004). This was further extended to the coupling of fluorescent-labels or -proteins to proteins of interest (Matsumoto et al, 2012, 2016; Ott et al, 2016) and the cross linking of enzymes (Li et al, 2017) or ligation of two protein domains (Omura et al, 2018; Raltchev et al, 2018). Peptide ligation to proteins has, among others, been exemplified by coupling peptides containing thioesters (Ling et al, 2012), cell penetrating ability (Van Lith et al, 2017), non-canonical AAs (Ke et al, 2017), antimicrobial activity (Touti et al, 2018), purification tags (Bellucci et al, 2013), isotopes for labeling (e.g., NMR) (Freiburger et al, 2015; Williams et al, 2016; Sonntag et al, 2017), or ligation handles such as azides (Ta et al, 2018).…”

Functional protein nanostructures: a chemical toolbox

“…In order to prepare protein dimers or homoprotein polymers, a single site of the protein is usually modified 27,28. In contrast, the formation of higher ordered structures such as rings or nanotubes requires the introduction of two or more recognition motif at the protein surface 29,30. The most straightforward strategy is the chemical modification of native proteins at already available single amino acid residues such as cysteines, disulfides, or amines.…”

“…4d). 86 To confer additional self-assembling handles on streptavidin, a twigged streptavidin polymer was engineered by Tanaka and co-workers as a scaffold for hetero-protein assembly 30. A sortase A recognition site and a horseradish peroxidase recognition site were genetically incorporated into the N- and C-termini of streptavidin, respectively, that allowed the immobilization of two different proteins via biotin–streptavidin interaction and sortase A-mediated ligation 30…”