Cold-active protease CP70

Details Cold-active protease CP70 Cold-active protease CP70

1998-07-27

2001-03-13

Saidha, Tekchand (Department: 1652)

Chemistry: molecular biology and microbiology

Enzyme , proenzyme; compositions thereof; process for...

Hydrolase

C435S219000, C435S850000, C530S300000, C530S412000

Reexamination Certificate

active

06200793

ABSTRACT:

BACKGROUND OF THE INVENTION
(i) Field of the Invention
The present invention relates to a protease having a high activity at a low temperature and its utilization.
(ii) Description of the Related Art
Psychrophilic bacteria have been known for a long time, and their existence can be confirmed extensively in low temperature circumstances. For example, psychrophilic bacteria can be isolated from soils, fishery products, milk products as well as artificial low temperature circumstances. Researches on psychrophilic bacteria have been conducted in accordance with food microbiological requirements, but they have principally been confined to those with respect to the phylogeny of microorganisms.
Enzymes obtained from psychrophilic bacteria are expected to be cold-active is enzymes having an optimal temperature in a low temperature range. The cold-active enzyme which acts efficiently at the low temperature is considered to be capable of being incorporated into, for example, a detergent which can be used even in water at the low temperature. It is also considered to be employed for a chemical reaction in the presence of an organic solvent which is volatile at ordinary temperature and for the quality improvement of foods at the low temperature at which the foods do not rot. Furthermore, the investigation on the enzyme derived from the psychrophilic bacteria is fairly interesting to reveal the physiological functions and the adaptation mechanism to the low temperature of the psychrophilic bacteria.
SUMMARY OF THE INVENTION
The present inventors have now found that a protease can be isolated from the supernatant liquid of a culture medium of a
Flavobacterium balustinum
P104 strain and then purified, and that the isolated and purified protease has activity at a low temperature. The present invention is based on such knowledge.
Therefore, an object of the present invention is to provide a cold-active protease.
Another object of the present invention is to provide a method for preparing the above-mentioned cold-active protease by the use of a
Flavobacterium balustinum
P104 strain.
Still another object of the present invention is to provide a peptide comprising an amino acid sequence present at the N terminal of the cold-active protease.
The protease according to the present invention has a part or all of the following physicochemical properties.
Specific activity and substrate specificity: The protease acts on casein, gelatin, hemoglobin and albumin to specifically decompose them in the order of casein, gelatin, hemoglobin and albumin.
Optimal pH: 8.0
pH stability: The protease is stable in the range of pH 6.5 to pH 10.0 at 30° C. for 1 hour.
The protease according to the present invention further has a part or all of the following physicochemical properties.
Optimal temperature: About 40° C.
Temperature stability: At pH 7 for 1 hour, the protease is scarcely inactivated at a temperature up to 30° C., but it is inactivated at 40° C. as much as about 40% and completely inactivated at 50° C. in about 10 minutes.
Enzyme activity: The protease has about 50% or more of its maximum activity at 20° C.
The active center of the enzyme is serine.
The molecular weight of the protease is about 70 kDa as measured by SDS-PAGE.
Furthermore, the protease according to the present invention consists of a protein containing a part or all of an amino acid sequence described in SEQ ID NO:1, or a protein containing a part or all of an amino acid sequence described in SEQ ID NO:1 at its N terminal.
According to another aspect of the present invention, we provide a protease having about 50% or more of its maximum activity at 20° C.
According to still other aspects of the present invention, we provide a protease which consists of a protein containing a part or all of an amino acid sequence described in SEQ ID NO:1, and a protease which consists of a protein containing a part or all of an amino acid sequence described in SEQ ID NO: 1 at its N terminal.
A method for preparing the above-mentioned protease according to the present invention comprises the steps of culturing
Flavobacterium balustinum
P104 (FERM BP-5006) for producing tprotease, and then collecting the protease from its culture medium.


REFERENCES:
patent: 4778878 (1988-10-01), Adams et al.
patent: 5646028 (1997-07-01), Leigh
Kato et al., “Protease Formation by a Marine Psychrophilic Bacterium.”: Agr. Biol. Chem., vol. 36, No. 7, p. 1177-1184, 1972.
Shikata et al., “Detection of Large COOH-Terminal Domains Processed from the Precursor ofSerratia marcescensSerine Protease in teh Outer Membrane ofEscherichia coli”, J. Biochem 111,627-632 (1992).
Kato et al., “Purification and Properties of Proteases from a Marine-psychrophilic Bacterium”, Agr. Biol. Chem., vol., No. 7 p. 1183-1192 (1972).

Affiliated with

Also associated with

Rating

Cold-active protease CP70 does not yet have a rating. At this time, there are no reviews or comments for this patent.

If you have personal experience with Cold-active protease CP70, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Cold-active protease CP70 will most certainly appreciate the feedback.

Rate now

Comments { 0 }

Profile ID: LFUS-PAI-O-2517805