# US 2014/0193890 A1 · Thermostable enzymes and methods of making and using the same > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 2014/0193890 A1 - Document ID: US20140193890A1 - Status: Published application - Kind: A1 - Application number: 13/991,605 - Filed: 2012-04-14 - Published: 2014-07-10 - Earliest claimed: 2012-01-31 - Assignees: University of Calcutta - Inventors: Arka Mukhopadhyay; Hirak Kumar Patra; Krishanu Chakraborti; Anjan Kr. Dasgupta; Dhrubajyoti Chattopadhyay - Web page: https://patentgrep.com/patent/US20140193890A1 ## CPC - C12N 9/96: Stabilising an enzyme by forming an adduct or a composition; Forming enzyme conjugates - A23K 20/189: Enzymes - A23L 5/25: {using enzymes} - C11D 3/386: Preparations containing enzymes {, e.g. protease or amylase} - C11D 7/16: Phosphates including polyphosphates - C11D 17/06: Powder; Flakes; Free-flowing mixtures; Sheets - C12N 9/88: Lyases (4.) - C12P 19/00: Preparation of compounds containing saccharide radicals (ketoaldonic acids C12P7/58) - C12Y 402/02002: Pectate lyase (4.2.2.2) - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) - A23K: FODDER - A23L: FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF (preservation thereof A23B) - C11D: DETERGENT COMPOSITIONS; USE OF SINGLE SUBSTANCES AS DETERGENTS; SOAP OR SOAP-MAKING; RESIN SOAPS; RECOVERY OF GLYCEROL - C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)} - C12Y: ENZYMES ## Abstract Provided herein are compositions and methods for enhancing enzyme activity, half-life and/or thermostability. Also provided herein are compositions and methods including the enhanced enzymes. Also provided herein are methods and compositions related to improved pectinolytic enzymes, such as pectate lyase, which exhibit enhanced activity, thermostability and/or longer half-life. ## Claims ### Claim 1 A composition, comprising: at least one nanoparticle, comprising an apatite moiety; and at least one enzyme in contact with the nanoparticle, the apatite moiety, or both, wherein the enzyme comprises a pectinolytic enzyme. ### Claim 2 (depends on 1) The composition of claim 1, wherein the apatite comprises hydroxyapatite. ### Claim 3 (canceled) ### Claim 4 (depends on 1) The composition of claim 1, wherein the enzyme comprises a pectate lyase, pectin lyase, polygalacturonase, polygalacturonate lyase, pectinesterase, pectinase, endopectate lyase, or combinations thereof. ### Claim 5 (depends on 1) The composition of claim 1, wherein the enzyme is a pectate lyase, pectin lyase, polygalacturonase, polygalacturonate lyase, pectinesterase, pectinase, endopectate lyase, or combinations thereof. ### Claim 6 (depends on 1) The composition of claim 1, wherein the enzyme is purified from one or more of Bacillus sp., Aspergillus sp., Penicillium sp., Sclerotinia sp., Stereum sp., Erwinia sp., Amycolata sp., Yersinia sp., Fusarium sp., Pseudomonas sp., Streptomyces sp., Candida sp., Rhodotorula sp., and Aureobasidium sp. ### Claim 7 (depends on 1) The composition of claim 1, wherein the enzyme comprises a recombinant enzyme. ### Claim 8 (depends on 1) The composition of claim 1, further comprising at least one divalent cation. ### Claim 9 (depends on 8) The composition of claim 8, wherein the divalent cation comprises calcium. ### Claim 10 (depends on 1) The composition of claim 1, wherein the enzyme in the composition exhibits one or more of the following characteristics: has an increased activity as compared to the same pectinolytic enzyme that is not in contact with the nanoparticle, the apatite moiety, or both; is more thermostable as compared to the same pectinolytic enzyme that is not in contact with the nanoparticle, the apatite moiety, or both; has an increased activity at a higher temperature as compared to the same pectinolytic enzyme that is not in contact with the nanoparticle, the apatite moiety, or both; has an increased activity at a higher temperature and is more thermostable as compared to the same pectinolytic enzyme that is not in contact with the nanoparticle, the apatite moiety, or both; has a longer half-life as compared to the same pectinolytic enzyme that is not in contact with the nanoparticle, the apatite moiety, or both. ### Claim at position 11 -14. (canceled) ### Claim 15 A method of making a thermostable enzyme composition, the method comprising: combining to form a mixture: a) a plurality of nanoparticles, wherein at least some of the nanoparticles comprise an apatite moiety; and b) at least one enzyme; under conditions in which the enzyme is in contact with the nanoparticles, the apatite moiety, or both. ### Claim 16 (depends on 15) The method of claim 15, wherein the apatite comprises hydroxyapatite. ### Claim 17 (depends on 15) The method of claim 15, wherein the enzyme is a pectate lyase. ### Claim 18 (depends on 15) The method of claim 15, further comprising combining one or more divalent cations into the mixture, wherein at least one of the divalent cations is selected from the group consisting of Ca2+, Mg2+, Mn2+, Co2+, and Cd2+. ### Claim at position 16 -24. (canceled) ### Claim 25 A method of treating a pectin-containing material, the method comprising: contacting the material with a composition comprising: a) at least one nanoparticle, comprising an apatite moiety; and b) a pectinolytic enzyme in contact with the nanoparticle, the apatite moiety, or both, for a time and under conditions wherein at least some of the pectin in the material is cleaved by the enzyme. ### Claim 26 (depends on 25) The method of claim 25, wherein the pectin-containing material comprises a textile, plant, detergent, biocomposite, wastewater, paper, oil, animal feed, food, beverage, or combinations thereof. ### Claim 27 (depends on 25) The method of claim 25, wherein the contacting is carried out at a temperature greater than or equal to about 55° C. ### Claim 28 (depends on 25) The method of claim 25, wherein the pectin-containing material comprises a food, and wherein the contacting is carried out at a temperature greater than or equal to 90° C. ### Claim 29 (depends on 25) The method of claim 25, wherein the composition further comprises at least one divalent cation. ### Claim 30 (canceled) ### Claim 31 A kit comprising: a) an pectinolytic enzyme; b) a plurality of nanoparticles comprising an apatite moiety; and c) instructions for combining the enzyme and the nanoparticles to form an enzyme composition. ### Claim 32 (canceled) ### Claim 33 (depends on 31) The kit of claim 31, wherein the apatite moiety comprises hydroxyapatite.