A6487
Alpha-Lytic Protease M190A Mutant
Synonym(s):
Alpha-Lytic Protease M190A Mutant, ALP M190A
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About This Item
UNSPSC Code:
12352204
NACRES:
NA.54
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recombinant
expressed in E. coli
form
liquid
specific activity
≥0.05 U/mg
mol wt
19.8 kDa (mature form)
optimum pH
5.0(storage)
7.5(activity)
pI
9.69
shipped in
dry ice
Storage temp.
−70°C
General description
Alpha-lytic protease (aLP) is an alternative specificity protease for proteomics applications, whose wild-type (WT) version cleaves after T, A, S, and V residues. The M190A (Met190 → Ala190) mutant of aLP has different cleavage specificities, and cleaves after M, F, and L residues. Both the WT and M190A forms of aLP generate peptides of similar average length as trypsin.
In WT aLP, the methionines at positions 190 and 213 cause WT aTP to have its particular specificity toward peptide substrates with small hydrophobic side chains at the P1 position. The M190A mutation gives the resulting mutant M190A aLP the ability to cleave peptide substrates with large hydrophobic side chains at the P1 side chain with greater efficiency compared to WT aLP, in addition to accommodating peptides with small hydrophobic side chains as before. Other structural changes in M190A aLP compared to WT aLP have been discussed.
The activity of M190A aLP in the presence of various solution components is as follows:
In WT aLP, the methionines at positions 190 and 213 cause WT aTP to have its particular specificity toward peptide substrates with small hydrophobic side chains at the P1 position. The M190A mutation gives the resulting mutant M190A aLP the ability to cleave peptide substrates with large hydrophobic side chains at the P1 side chain with greater efficiency compared to WT aLP, in addition to accommodating peptides with small hydrophobic side chains as before. Other structural changes in M190A aLP compared to WT aLP have been discussed.
The activity of M190A aLP in the presence of various solution components is as follows:
- 0.1% sodium deoxycholate: ~1.4-fold enhanced activity
- 1.0% sodium deoxycholate: nearly full activity
- 0.1% SDS: ~40% activity
- 1.0% SDS: ~30% activity
- 1 M guanidine HCl: ~20% activity
- 4 M guanidine HCl: ~1% activity (essentially inactivated)
Unit Definition
mmols of pNA produced per min per mg protein from 0.5 mM N-succinyl-Ala-Ala-Pro-Phe-PNA at 25 °C in 100 mM Tris-HCl (pH 7.5).
related product
Product No.
Description
Pricing
Storage Class
12 - Non Combustible Liquids
wgk_germany
WGK 1
flash_point_f
Not applicable
flash_point_c
Not applicable
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J E Mace et al.
Journal of molecular biology, 254(4), 720-736 (1995-12-08)
Gly216 in the active site of the broadly specific MA190 mutant of alpha-lytic protease has been found to be remarkably tolerant of amino acid substitutions. Side-chains as large as Trp can be accommodated within the substrate-binding pocket without abolishing catalysis
D W Miller et al.
Journal of molecular biology, 286(1), 267-278 (1999-02-05)
We have used alpha-lytic protease as a model system for exploring the relationship between the internal dynamics of an enzyme and its substrate specificity. The wild-type enzyme is highly specific for small substrates in its primary specificity pocket, while the
R Bone et al.
Nature, 339(6221), 191-195 (1989-05-18)
The substrate specificity of alpha-lytic protease has been changed dramatically, with a concomitant increase in activity, by replacing an active-site Met with Ala. The substrate specificity of both this mutant and another similar mutant are extraordinarily broad. X-ray crystallographic analysis
The α-Lytic Protease.
Whitaker, D.R.
Methods in Enzymology, 19, 599-613 (1970)
Jesse G Meyer et al.
Molecular & cellular proteomics : MCP, 13(3), 823-835 (2014-01-16)
Bottom-up proteomics studies traditionally involve proteome digestion with a single protease, trypsin. However, trypsin alone does not generate peptides that encompass the entire proteome. Alternative proteases have been explored, but most have specificity for charged amino acid side chains. Therefore
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