Phospholipases, nucleic acids encoding them and methods for making and using them

A phospholipase, nucleic acid technology, applied in the field of polynucleotides, can solve problems such as yield loss and quality decline

Inactive Publication Date: 2005-08-24
VERENIUM CORP (US)
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The neutralization process is very effective at removing free fatty acids and phos

Method used

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  • Phospholipases, nucleic acids encoding them and methods for making and using them
  • Phospholipases, nucleic acids encoding them and methods for making and using them
  • Phospholipases, nucleic acids encoding them and methods for making and using them

Examples

Experimental program
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Effect test

Embodiment 1

[0513] Example 1: BLAST program for sequence identity exploration

[0514] This example illustrates a typical sequence identity program used to determine whether a nucleic acid is within the scope of the invention. The NCBI BLAST 2.2.2 program was applied, with to blastp selected by default. Except for the default filtering setting, all default values ​​apply (ie, all parameters are set to default except for filtering set to OFF); in this position the "FF" setting is applied, which disables filtering. Due to the short length of the sequences, applying default filtering often leads to Karlin-Altschul violations. Default values ​​used in this example:

[0515] "Filter for low complexity: ON "Low complexity filtering: ON

[0516] >Word Size: 3 >Word Length: 3

[0517] >Matrix: Blosum62 >Matrix: Blosum 62

[0518] >Gap Costs: Existence: ll >Gap Costs: Existence: ll

[0519] >Extension: 1” >Extension: 1”

[0520] Other default settings are: low complexity filtering off, fo...

Embodiment 2

[0522] Example 2: Simulation of PLC-mediated degumming

[0523] This example illustrates the simulation of phospholipase C (PLC) mediated degumming.

[0524] Due to the poor solubility of phosphatidylcholine (PC) in water, it was initially dissolved in ethanol (100 mg / ml). For initial assays, stock solutions of PC were prepared by dissolving in 50 mM morpholinopropanesulfonic acid or 60 mM citric acid / NaOH at pH 6. PC stock solution (10 μl, 1 μg / μl) was added to 500 μl refined soybean oil (with 2% water) in an Eppendorf tube. To obtain an emulsion, the contents of the tube were mixed by vortex mixer for 3 minutes (see Figure 5 A). The oil and water phases were separated by centrifugation at 13,000 rpm for 1 minute ( Figure 5 B). The reaction tube was pre-incubated at the desired temperature (37°C, 50°C, or 60°C), and 3 μl of PLC (0.9 U / μl) from Bacillus cereus was added to the aqueous phase ( Figure 5 C). Disappearance of PC was analyzed by TLC using chloroform / met...

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Abstract

The present invention provides novel polypeptides having phospholipase activity, including, for example, phospholipase A, B, C and D activities, potato tuber storage protein activity, fatty acyl hydrolase (LAH) activity, nucleic acids encoding them and antibodies that bind them . Industrial processes involving the use of these phospholipases, such as oil degumming, and products involving the use of these phospholipases are also provided.

Description

field of invention [0001] The present invention generally relates to phospholipases, polynucleotides encoding phospholipases, methods of making and using these polynucleotides and polypeptides. In particular, the present invention provides novel polypeptides having phospholipase activity, nucleic acids encoding them, and antibodies that bind them. Industrial processes and products involving the use of these phospholipases are also provided. Background technique [0002] Phospholipases are enzymes that hydrolyze the ester bonds of phospholipids. Corresponding to their importance in phospholipid metabolism, these enzymes are widespread in eukaryotes and prokaryotes. Phospholipases affect metabolism, formation and reorganization of biofilms, and phospholipases are also involved in signaling cascades. Several types of phospholipases are known which differ in specificity according to the position of the bond attacked by the phospholipid molecule. Phospholipase A1 (PLA1) remov...

Claims

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Application Information

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IPC IPC(8): A01H5/00A01K67/027C07K7/00C07K14/00C07K16/40C07K19/00C12N1/15C12N1/19C12N1/21C12N5/10C12N9/16C12N11/00C12N15/09C12P7/02C12P21/08C12Q1/02C12Q1/34C12Q1/68G01N33/53G01N37/00
CPCC12N9/18Y02E50/13A01K2217/05C12N9/16C07K2319/00
Inventor S·格拉马蒂科瓦G·黑尔伍德D·拉姆N·巴顿
Owner VERENIUM CORP (US)
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