Evaluation of resins for on-bead screening: A study of papain and chymotrypsin specificity using PEGA-bound combinatorial peptide libraries

Bioorganic and Medicinal Chemistry Letters

Volumn 8, Issue 21, 1998, Pages 2997-3002

  Leon, Susanna ^a   Quarrell, Rachel ^a   Lowe, Gordon ^a  

^a UNIVERSITY OF OXFORD   (United Kingdom)

Author keywords

[No Author keywords available]

Indexed keywords

CHYMOTRYPSIN; PAPAIN; RESIN;

ARTICLE; ENZYMATIC ASSAY; ENZYME SPECIFICITY; PEPTIDE ANALYSIS; SCREENING;

CHYMOTRYPSIN; PAPAIN; PEPTIDES; POLYETHYLENE GLYCOLS; RESINS, PLANT;

EID: 0032211605     PISSN: 0960894X     EISSN: None     Source Type: Journal    
DOI: 10.1016/S0960-894X(98)00534-4     Document Type: Article

References (29)

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8. max,em 525 nm) and the fluorescence intensity of partially-labelled dansyl-beads showed an approximately linear dependence in relation to loading. Spectra were obtained fluorimetrically by stirring a suspension of beads in DMF in a cuvette.
9. 9. Edman sequencing results: Cycle 1: L (44), G (32); Cycle 2: G (44); Cycle 3: G (25). Figures in brackets denote yields in pmol/bead, and are average values calculated from duplicate analyses of single bead samples. PEGA beads of a diameter range 150-300 μm were used, with a peptide loading of 60-180 pmol/bead as determined by Edman sequencing. Prolonged treatment of the beads with papain showed quantitative levels of cleavage.
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13. -4 M would not be capable of drawing the enzyme into the bead against the significant barrier.
14. 14. Edman sequencing results: Cycle 1: G (4.2), L (1.7), F (1.6); Cycle 2: G (4.3), F (2.4); Cycle 3: G (3.5), F (1.5); Cycle 4: G (2.7), L (1.7); Cycle 5: G (2.8), L (1.3); Cycle 6 (2.6). Figures in brackets are average yields in pmol/bead, calculated from the duplicate analyses of samples comprising 13 beads. Beads were of mean diameter 176 μm, with an original peptide loading of approx. 140 pmol/bead determined by Edman sequencing.
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24. 24. The assays with library (2) were limited to 300 sequences due to a logistical problem during synthesis.
25. 25. Edman sequencing results: Sample 1: Cycle 1: G (20); Cycle 2: T (4); Cycle 3: G (18); Cycle 4: G (16). Sample 2: Cycle 1: G (20); Cycle 2: T (4); Cycle 3: G (16); Cycle 4: G (15). Sample 3: Cycle 1: G (84); Cycle 2: L (77); Cycle 3: G (69); Cycle 4: G (59). Sample 4: Cycle 1: G (13); Cycle 2: E (8); Cycle 3: G (9); Cycle 4: G (10). Low yields of threonine are inherent to the Edman process.
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