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US 2003/0144165 A1Published application
Lipolytic enzyme variant
AssigneeNovozymes A/S
InventorErwin Ludo Roggen
Life of this patent
Estimating the term…
Abstract
The properties of a fungal lipolytic enzyme can be altered by substituting amino acid residues corresponding to certain specified amino acid residues in the T. lanuginosus lipase. The altered property may be, e.g., an increased thermostability, an altered pH dependence, or an altered substrate specificity.
Claims18
A lipolytic enzyme which is a variant of a parent fungal lipolytic enzyme, comprising one or more amino acid substitutions corresponding to any of the following in SEQ ID NO:
- 1:
- Q15 to A, C, D, E, F, G, I, K, L, M, N, P. R, S, T, V, W, Y;
- Y16 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W;
- A18 to C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W, Y;
- A19 to C, D, E, F, G, H, I, K, L, M, N, Q, R, S, V, W, Y;
- A20 to C, D, E, F, G, H, I, K, L, M, N, Q, R, S, T, V, W, Y;
- N25 to A, C, D, E, F, G, H, I, K, L, M, P, Q, R, S, T, V, W, Y;
- N26 to A, D, E, F, G, H, I, K, L, M, P, Q, R, S, T, V, W, Y;
- E43 to A, C, D, F, G, H, I, K, L, M, N, R, S, T, V, W, Y;
- V44 to A, C, D, E, F, G, H, I, K, L, M, N, Q, R, S, T, W, Y;
- K46 to A, C, D, E, F, G, H, I, L, M, N, Q, S, T, V, W, Y;
- A47 to C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, W, Y;
- A49 to C, D, E, F, G, H, I, K, L, M, N, Q, R, S, V, W, Y;
- L52 to A, C, D, E, F, G, H, I, K, M, N, P, Q, R, S, T, V, W, Y;
- Y53 to A, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W;
- S54 to A, C, D, E, F, G, H, I, K, L, M, N, Q, R, T, V, W, Y;
- G65 to A, C, D, E, F, H, I, K, L, M, N, P, Q, R, S, T, V, W, Y;
- L67 to A, C, D, E, F, G, H, I, K, M, N, Q, R, S, T, V, W, Y;
- A68 to C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W, Y;
- L69 to A, C, D, E, F, G, H, I, K, M, N, P, Q, S, T, V, W, Y;
- T72 to A, C, D, E, F, G, H, I, L, M, N, P, Q, R, S, V, W, Y;
- K74 to A, C, D, E, F, G, H, I, L, M, N, P, Q, R, S, T, V, W, Y;
- L75 to A, C, D, E, F, G, H, I, K, M, N, P, Q, R, S, T, V, W, Y;
- V77 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, W, Y;
- S79 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, W, Y;
- R81 to A, D, E, F, G, H, I, K, L, M, N, P, Q, S, T, V, W, Y;
- S83 to A, C, D, E, F, G, H, I, K, L, M, N, Q, R, V, W, Y;
- S85 to A, D, E, G, H, I, L, M, N. Q, V, W, Y;
- W89 to A, C, D, E, F, H, I, K, L, M, N, P, Q, R, S, T, V, Y;
- L97 to A, C, D, E, F, G, H, I, K, N, P, R, S, T, W, Y;
- K98 to A, C, G, H, L, M, N, P, Q, S, T, V, W, Y;
- E99 to C, F, G, I, M, P, W, Y;
- G106 to A, C, D, E, F, H, I, K, L, M, N, P, Q, R, T, V, W, Y;
- C107 to A, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W, Y;
- R108 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, S, T, V, Y;
- G109 to A, C, D, E, F, H, I, K, L, M, N, P, Q, R, S, T, W, Y;
- T123 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, V, W, Y;
- L124 to A, C, D, E, F, G, H, I, K, M, N, P, Q, R, T. V, W, Y;
- K127 to A, D, E, F, G, H, I, L, M, N, P, Q, R, S, T, V, W, Y;
- E129 to A, C, D, F, G, H, I, L, M, N, P, Q, R, S, T, V, W, Y;
- A131 to C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W, Y;
- V132 to A, C, D, E, F, G, H, I, K, L, N, P, Q, R, S, T, W, Y;
- Y138 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W;
- V140 to A, C, D, E, F, G, H, I, K, L, M, N, P, R, S, T, W, Y;
- L147 to A, C, D, E, F, G, H, I, K, M, N, P, Q, R, S, T, V, W, Y;
- A150 to C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W, Y;
- T153 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, V, W, Y;
- Y164 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W;
- D165 to A, C, E, F, G, H, I, K, L, M, N, Q, S, T, V, W, Y;
- D167 to A, C, E, F, H, K, L, M, N, P, Q, S, T, V, W, Y;
- S170 to A, C, D, E, F, G, H, I, K, L, M, N, Q, R, T, V, W, Y;
- Y171 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W;
- G172 to A, C, D, E, F, H, I, K, L, M, N, P, Q, R, S, T, V, W, Y;
- A173 to C, D, E, F, G, H, I, K, L, M, N, Q, R, S, T, V, W, Y;
- P174 to A, C, D, E, F, G, H, I, K, L, M, N, Q, R, S, T, V, W, Y;
- R175 to A, C, D, E, F, G, H, I, K, L, M, N, Q, S, T, V, W, Y;
- V176 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, W, Y;
- G177 to A, C, D, E, F, H, I, K, L, M, N, P, Q, R. S, T, V, W, Y;
- R179 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, S, T, V, W, Y;
- A182 to C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W. Y;
- Y194 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W;
- H198 to A, C, D, E, F, G, H, K, L, M , P, Q, R. S, T, V, W, Y;
- N200 to A, C, D, E, F, G, H, I, K, L, M, P, Q, S, T, V, W, Y;
- P207 to A, C, D, E, F, G, H, I, K, L, M, N, Q, R, S, T, V, W, Y;
- P208 to A, C, D, E, F. G, H, I, K, L, M, N, Q, R, S, T, V, W, Y;
- R209 to C, D, F, G, H, I, K, L, M, N, Q, T, V, W, Y;
- G212 to A, C, D, E, F, H, I, K, L, M, N, P, Q, R, S, T, V, W, Y;
- S214 to A, C, D, E, F, G. H, I, K, L, M, N, P, Q, R, T, V, W, Y;
- H215 to A, C, D, E, F, G, I, K, L, M, N, P, Q, R, S. T, V, W, Y;
- S216 to A, C, D, E, F, G, H, I, K, L, M, N, Q, R, T, V, W, Y;
- S217 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, T, V, W, Y;
- P218 to A, C, D, E, F, G, H, I, K, L, M, N, Q, R, S, T, V, W, Y;
- E219 to C, D, F, H, I, M, P, W, Y;
- Y220 to A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W;
- K223 to A, C, D, E, F, G, H, I, L, M, N, Q, S, T, V, W, Y;
- S224 to A, C, D, E, F, G, H, I, K, L, M, N, Q, T, V, W, Y;
- D234 to C, E, F, H, I, M, W;
- I235 to A, C, D, E, F, G, H, K, L, M, N, P, Q, R, S, T, V, W, Y;
- K237 to A, C, D, E, F, G, H, L, N, P, Q, S, T, V, W, Y;
- I238 to A, C, D, E, F, G, H, K, L, M, N, P, Q, R, S, T, V, W, Y;
- D242 to C, E, F, G, H, I, M, P, W, Y;
- A243 to C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, V, W, Y;
- P250 to A, C, D, E, F, G, H, I, K, L, M, N, Q, R, S, T, V, W, Y;
- P253 to A, C, D, E, F, G, H, I, K, L, M, N, Q, S, T, V, W, Y;
- D254 to C, E, F, H, I, M, P, Y;
- I255 to C, D, E, F, H, L, M, N, Q, W, Y;
- P256 to C, E, F, G, H, I, K, L, M, N, Q, R, V, W, Y;
- Y261 to A, C, E, F, G, H, L, M, N, P, Q, R, S, T, V.
85 more elements
The lipolytic enzyme of any preceding claim wherein the parent lipolytic enzyme has at least 80% homology with SEQ ID NO:
- 1.
The lipolytic enzyme of the preceding claim wherein the parent lipolytic enzyme is the lipase produced by Thermomyces lanuginosus DSM 4109 and having the amino acid sequence of SEQ ID NO:
- 1.
The lipolytic enzyme of any preceding claim which has one, two, three, four, five or six of said substitutions.
A DNA sequence encoding the lipolytic enzyme of any preceding claim.
A vector comprising the DNA sequence of the preceding claim.
A method of producing a lipolytic enzyme comprising:
- a) selecting a parent polypeptide which is a fungal lipolytic enzyme,
- b) preparing a polypeptide derived from the parent polypeptide by substituting at least one amino acid residue corresponding to any of the following amino acids in SEQ ID NO:
- 1 and optionally substituting one or more other amino acids:
- Q15, Y16, A18, A19, A20, N25, N26, E43, V44, K46, A47, A49, L52, Y53, S54, G65, L67, A68, L69, T72, K74, L75, V77, S79, R81, S83, S85, W89, D96, L97, K98, E99, G106, C107, R108, G109, T123, L124, K127, E129, A131, V132, Y138, V140, L147, A150, T153, Y164, D165, D167, S170, Y171, G172, A173, P174, R175, V176, G177, R179, A182, Y194, H198, N200, P207, P208, R209, G212, S214, H215, S216, S217, P218, E219, Y220, K223, S224, D234, I235, K237, I238, D242 to A243, P250, P253, D254, I255, P256, Y261,
- c) testing the lipolytic enzyme activity of the polypeptide,
- d) selecting a polypeptide having lipolytic enzyme activity and an altered property compared to the parent polypeptide, and
- e) producing the selected polypeptide.
4 more elements
The method of the preceding claim wherein the parent polypeptide has at least 80% homology with SEQ ID NO:
- 1.
The method of the preceding claim wherein the parent polypeptide is the lipase produced by Thermomyces lanuginosus DSM 4109 and having the amino acid sequence of SEQ ID NO:
- 1.
Dependencies are inferred from the claim text.