Methods in Molecular Biology • 16 Enzymes of Molecular Biology

Peptide Production 295 morpholine acetate (pH 8.5). For pepsin, 5% acetic acid or 10 mM HC1 is used. Ammonium acetate (0.1M) can ...

296 Sweeney and Walker proteolytic fragments from the enzyme being used. Bacterial growth and protein cleavage by released enzym ...

Peptide Production 297 then taken, a final addition of enzyme made, and incubation continued for a further 8 h, when a final sam ...

298 Sweeney and Walker 3.6.2. Clostripain Note that the enzyme may need to be activated prior to use (see Section 2.2.7.). Disso ...

Peptide Production 299 3.6.10. Trypsin Dissolve the substrate in 0.1Mammonium bicarbonate. Add enzyme to 1-2% (w/w), and incubat ...

300 Sweeney and Walker Labouesse, B. and Gros, P. (1960) Laclostripaine, prot6ase de Clostridium histolyticum Purification et a ...

Peptide Production 301 Schenkein, I., Franklin, E. C., and Frangione, B. (1981) Proteolytic enzymes from the mouse submaxillary ...

302 Sweeney and Walker Drapeau, G. R. (1977) Cleavage at glutamic acid with staphlococcal protease. Methods Enzymol. 47, 189-19 ...

Peptide Production 303 Poncz, L. and Dearborn, D. G. (1983) The resistance to tryptic hydrolysis of peptide bonds adjacent to N ...

CHAPTER 16 Proteinase K (EC 3.4.21.14) Patricia J. Sweeney and John M. Walker Introduction Proteinase K is a serine protease an ...

306 Sweeney and Walker method (8-10). Because of its broad specificity and its ability to func- tion in the presence of detergen ...

Proteinase K 307 2.5. Assay The assay is based on the hydrolysis ofN-acetyl-L-tyrosine ethylester (ATEE). One unit will hydrolyz ...

308 Sweeney and Walker Stock solutions of the enzyme are normally prepared in water at 10- 25 mg/mL and stored at -20°C. However ...

Proteinase K 309 However, misleading results can be obtained if exposed regions of the polypeptide are inherently protease resis ...

310 Sweeney and Walker McGookin, R. (1984) RNA extraction by the proteinase K method, in Methods in Molecular Biology, vol. 2: ...

Proteinase K 311 P~ihler, A., Banerjee, A., Dattagupta, J. K., Fujiwana, T., Lindner, K., Pal, G. P., Suck, D., Weber, G., and ...

CHAPTER 17 Carboxypeptidase Y (EC 3.4.16.1) Julia S. Winder and John M. Walker Introduction Carboxypeptidases are proteolytic e ...

314 Winder and Walker hydroxyproline, or lysine residues (1). The specificity of carboxypep- tidase B is far more restricted, cl ...

Carboxypeptidase Y 315 2.3. Physical and Chemical Properties CPY has a mol mass of 61,000-63,000 Da as determined by SDS gel ele ...

316 Winder and Walker Denaturing Agents Eighty percent of activity remains after incubation with 6M urea at 25°C for 1 h (3). ...