Protein polymer modification

The improvement of its activity and stability has been approach by the use of GE tools (see Refs. [398] and [399], respectively). A process drawback is the fact that the oxidation of hydrophobic compounds in an organic solvent becomes limited by substrate partition between the active site of the enzyme and the bulk solvent [398], To provide the biocatalyst soluble with a hydrophobic active site access, keeping its solubility in organic solvents, a double chemical modification on horse heart cytochrome c has been performed [400,401], First, to increase the active-site hydrophobicity, a methyl esterification on the heme propionates was performed. Then, polyethylene glycol (PEG) was used for a surface modification of the protein, yielding a protein-polymer conjugates that are soluble in organic solvents. 

Another approach was recently developed by Smeenk et al. [90]. They used the combination of protein engineering and polymer modification for the creation of a series of silk-based block copolymers. Spider silk consists of two major domains, a j0-sheet crystalline domain, which gives strength to the protein, and a less well-defined amorphous domain, which introduces elasticity and toughness. Smeenk and co-workers produced a j0-sheet forming protein... 

Modification of the protein amine groups is the procedure most frequently used to produce antibody-liposome conjugates. Early procedures used crosslinking agents, such as l-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) (27,28) in the presence of preformed liposomes containing a lipophilic carboxylic acid. Condensing agents like these tend to form protein-protein polymers. Control of these reactions is typically difficult and complex, and as a result separation of the liposomes from protein polymers is a... 

FIG. 13 Log-log plot of the binding isotherms of amphiphilic proteins to amphiphilic poly anions. ( , o) /3-lactoglobulin to copolymer of maleic acid and octyl- or dodecyl vinylether, respectively pH 8.7 and ionic strength 0.1 M (data from Ref. 50). (0, , A) bovine serum albumin to C18 modified poly(acrylic acid), pH 9.0 in 30 mM NaOH-boric acid buffer. Modification rate 10, 3, and 1 mol% respectively (data from Ref. 63). Free protein in /rniol/L protein/polymer ratio in number of protein globules per 1000 monomers. 

Non-natural amino acids can be incorporated into peptides and polypeptides via several different methodologies. Solid-phase peptide synthesis (SPPS) is a straightforward method for incorporation of non-natural amino acids and allows the incorporation of essentially any amino acid but is limited by the size of the peptides produced 18). Suppression-based strategies, both in vitro and in vivo, have been developed for site specific incorporation of diverse set non-natural amino acids into natural and synthetic polypeptides 19). Alternatively, auxotrophic expression hosts have been used for multisite incorporation of nonnatural amino acid in protein polymers, where multiple natural amino acids of one type can be replaced with non-natural analogues during protein biosynthesis (20, 21). Multisite incorporation of non-natural amino acids in the synthesis of protein polymeric materials facilitates chemical modification at multiple sites and can modulate the physical properties of the protein polymers (22). 

The capacity of pathogenic bacteria to adhere to mucosal membranes has been exploited in the modification of new mucoadhesive polymers. The ability of bacteria to adhere to a specific target is rooted from particular cell-surface components or appendages, known as fimbriae, which promote adhesion to other cells or inanimate surfaces. Fimbriae are extracellular, long thread-like protein polymers of bacteria that play a major role in many diseases. It has been reported that Escherichia coli adheres specifically to the lymphoid follicle epithelium of the ileal Peyer s patch in rabbits. Similarly, different staphylococci possess the ability to adhere specifically to the surface of mucus gel layers and not mucus-free surfaces. Thus, polymers have been modified by the attachment of these fimbriae to enhance mucoadhesion. An attachment protein derived from E. coli, K99-fimbriae, has been covalently attached to polyacrylic acid networks in an attempt to provide a novel polymer with enhanced adhesive properties (Figure 52.7). ... 

ATRP initiators or RAFT CTAs containing protein-reactive moieties can be used to yield a-functionalized polymers, while postpolymerization modifications can be performed to yield ffi-ftmctionalized polymers. Figure 3 contains examples of functionalized ATRP initiators and RAFT CTAs. The resulting polymers can then be used to produce protein-polymer conjugates consisting of a single polymer chain and a single protein. These techniques can also be combined to synthesize polymer-linked protein dimers. 

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