Thiol amino acid sequences

To ascertain the upper limit of protein thermostability and to evaluate the effect of additional disulfide bridges on the enhancement of protein thermostability, additional cysteine residues were introduced into several unrelated proteins by site-directed mutagenesis and deactivation behavior tested at 100°C (Volkin, 1987). All the proteins investigated underwent heat-induced beta-elimination of cystine residues in the pH 4—8 range with first-order kinetics and similar deactivation constants kj that just depended on pH 0.8 0.3 h-1 at pH 8.0 and 0.06 0.02 h 1 at pH 6.0. These results indicate that beta-elimination is independent of both primary amino acid sequence and the presence of secondary structure elements. Elimination of disulfides produces free thiols that cause yet another deleterious reaction in proteins, heat-induced disulfide interchange, which can be much faster than beta-elimination. 

G, Kamphuis, J. Drentii, and E. N. Baker. Thiol protease. Comparative studies based on the high-resolution structures of papain and actinidin, and an amino acid sequence information far cathenaim B and R and stem bromelain. J. MoL BioL /62 317 (19B5). 

As shown in Figure 6, the similar properties that exist among the plant proteases can also be extended to the amino acid sequences which surround the active thiol group of these enzymes (68-73). 

Figure 6. Amino acid sequences around the active SH group of thiol proteases...

The complete amino acid sequence of papain is shown in Figure 10. The structure is based on the combined eflForts of several groups of investigators (69, 72, 79). Cysteine-25 has been identified as the active thiol... 

Figure 10.9. Amino acid sequences of cyclopeptides 1 and 2. each attached via three cysteine thiol groups to gold surfaces [99],...

Wintersberger2 isolated from bovine pancreatic carboxypeptidase a peptide containing a thiol group involved in the binding of zinc. He first removed the zinc by exposure to a chelating agent (1,10-phenanthroline) or by denaturation, and then labeled the reactive sulfhydryl group by reaction with DDPM and determined the amino acid sequence. 

N-(2-Hydroxypropyl)methacrylamide copolymers bearing oligopeptide side chains with a terminal p-nitroaniline (NAp) were incubated with purified lysosomal enzymes (Tritosomes) in vitro, and cleavage of these chains was measured by monitoring the release of NAp. It was shown that certain sequences liberate the terminal residue on incubation at pH 5.5 with lysosomal enzymes Recently, the importance of lysosomal thiol-proteinases in N-(2-hydroxypropyl)methacrylamide copolymer side chain cleavage was discovered The synthesis of side-chain amino acid sequences chosen to match the known specificities of certain lysosomal thiol-proteinases resulted in a higher initial rate of NAp release and a greater extent of release (up to 50% of NAp bound for an incubation period of 5 h) Incubation of substrates in vitro... 

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