(12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization International Bureau (10) International Publication Number (43) International Publication Date WO 2014/100018 Al 26 June 2014 (26.06.2014) W P O P C T (51) International Patent Classification: (81) Designated States (unless otherwise indicated, for every C12N 9/24 (2006.01) kind of national protection available): AE, AG, AL, AM, AO, AT, AU, AZ, BA, BB, BG, BH, BN, BR, BW, BY, (21) International Application Number: BZ, CA, CH, CL, CN, CO, CR, CU, CZ, DE, DK, DM, PCT/US2013/075829 DO, DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, (22) International Filing Date: HN, HR, HU, ID, IL, IN, IR, IS, JP, KE, KG, KN, KP, KR, 17 December 2013 (17. 12.2013) KZ, LA, LC, LK, LR, LS, LT, LU, LY, MA, MD, ME, MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ, (25) Filing Language: English OM, PA, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SA, (26) Publication Language: English SC, SD, SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM, TN, TR, TT, TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, (30) Priority Data: ZW. 61/739,267 19 December 2012 (19. 12.2012) US (84) Designated States (unless otherwise indicated, for every (71) Applicant: DANISCO US INC. [US/US]; 925 Page Mill kind of regional protection available): ARIPO (BW, GH, Road, Palo Alto, California 94304 (US). GM, KE, LR, LS, MW, MZ, NA, RW, SD, SL, SZ, TZ, UG, ZM, ZW), Eurasian (AM, AZ, BY, KG, KZ, RU, TJ, (72) Inventors: BARNETT, Christopher C ; Danisco US Inc., TM), European (AL, AT, BE, BG, CH, CY, CZ, DE, DK, 925 Page Mill Road, Palo Alto, California 94304 (US). EE, ES, FI, FR, GB, GR, HR, HU, IE, IS, IT, LT, LU, LV, DEMASO, Connie; Danisco US Inc., 925 Page Mill Road, MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, SI, SK, SM, Palo Alto, California 94304 (US). DODGE, Eric; Danisco TR), OAPI (BF, BJ, CF, CG, CI, CM, GA, GN, GQ, GW, US Inc., 925 Page Mill Road, Palo Alto, California 94304 KM, ML, MR, NE, SN, TD, TG). (US). HARRIS, Kathleen A.; Danisco US Inc., 925 Page Mill Road, Palo Alto, California 94304 (US). QI, Rachel Published: B.; Danisco US Inc., 925 Page Mill Road, Palo Alto, Cali — with international search report (Art. 21(3)) fornia 94304 (US). (74) Agent: KONG, Lawrence B.; Danisco US Inc., 925 Page Mill Road, Palo Alto, California 94304 (US). (54) Title: NOVEL MANNANASE, COMPOSITIONS AND METHODS OF USE THEREOF < 00 o o Figure 6-1. Cleaning performance of Blemanl in the presence of protease. (57) Abstract: The present compositions and methods relate to an endo-beta-mannanase cloned from Bacillus lentus, polynuc - © leotides encoding the endo-beta-mannanase, and methods of use thereof. Formulations containing the endo-beta-mannanase are highly suitable for use as detergents. NOVEL MANNANASE, COMPOSITIONS AND METHODS OF USE THEREOF CROSS REFERENCE TO RELATED APPLICATIONS [001] This application claims benefit of priority from U.S. provisional patent application no. USSN 61/739,267, filed on 19 December 2012, and is incorporated herein by reference in its entirety. BACKGROUND [002] Current laundry detergent and fabric care compositions include a complex combination of active ingredients such as surfactants, enzymes (e.g., protease, amylase, lipase, mannanase, and/or cellulase), bleaching agents, a builder system, suds suppressors, soil-suspending agents, soil-release agents, optical brighteners, softening agents, dispersants, dye transfer inhibition compounds, abrasives, bactericides, and perfumes. [003] Mannanase enzymes have been employed in detergent cleaning compositions for the removal of gum stains by hydrolyzing mannans. However, enzymes are often inhibited by surfactants and other components present in cleaning compositions, which interferes with their ability to remove stains. For instance, proteases in laundry detergents may degrade mannanases before the removal of a gum stain. In addition, mannanases may have a limited pH and/or temperature range at which they are active, which may make them unsuitable for certain formulations and washing conditions. Accordingly, the need exists for mannanases that retain activity in the harsh environment of cleaning compositions. SUMMARY [004] The present compositions and methods relate to a mannanase cloned from Bacillus lentus (Blemanl). Formulations containing the endo- -mannanase are highly suitable for use as detergents. [005] In one embodiment, the invention is a recombinant polypeptide comprising a catalytic domain of an endo- -mannanase, wherein the catalytic domain is at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO:2. In other embodiments, the invention is a recombinant polypeptide comprising a mature from of an endo- -mannanase, wherein the mature form is at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO:2. In each embodiment, the polypeptide of the invention has mannanase activity. In some embodiments, the polypeptide of the invention has mannanase activity on an AZO-Carob Galactomannan or Locust Bean Gum substrate. [006] In some embodiments, the polypeptide of the invention has mannanase activity at pH values of between 5 and 11, or between 7 and 11, or between 7 and 9. In some embodiments, the polypeptide of the invention retains greater than 50% mannanase activity at pH values of between 5 and 11, or between 7 and 11, or between 7 and 9, or between 7 and 8. In some embodiments, the polypeptide of the invention retains greater than 75% mannanase activity at pH values of between 5 and 11, or between 7 and 11, or between 7 and 9, or between 7 and 8. In some embodiments, the polypeptide of the invention has mannanase activity at temperature values of between 10°C and 60°C, or between 20°C and 60°C, or between 30°C and 60°C, or between 40°C and 60°C. [007] In some embodiments, the polypeptide of the invention has measurable mannanase activity in the presence of detergent. In some embodiments, the polypeptide has measurable mannanase activity in the presence of a protease. In some embodiments, the polypeptide retains greater than 50% mannanase activity at pH values of between 5 and 11, or between 7 and 11, or between 7 and 9, or between 7 and 8 in the presence of protease. In some embodiments, the polypeptide of the invention retains greater than 75% mannanase activity at pH values of between 5 and 11, or between 7 and 11, or between 7 and 9, or between 7 and 8 in the presence of protease. In some embodiments, the polypeptide has mannanase activity at temperature values of between 10°C and 60°C, or between 20°C and 60°C, or between 30°C and 60°C, or between 40°C and 60°C in the presence of protease. In some embodiments, the polypeptide retains at least 50% mannanase activity at temperature values of between 10°C and 60°C, or between 20°C and 60°C, or between 30°C and 60°C, or between 40°C and 60°C in the presence of protease. In some embodiments, the polypeptide is capable of hydrolyzing a substrate selected from the group consisting of chocolate ice cream, chocolate pudding, guar gum, locust bean gum, and combinations thereof. In some embodiments, the polypeptide further comprises a native or non-native signal peptide. In some embodiments, the polypeptide further comprises at least one carbohydrate-binding module. In other embodiments, the polypeptide does not comprise a carbohydrate-binding module. [008] In some embodiments, the invention is a detergent composition having at least one recombinant polypeptide as described above. In some embodiments, the invention is a detergent composition having a polypeptide as described above, wherein the polypeptide is active at a temperature between 10°C and 60°C, or 20°C and 60°C, or 30°C and 60°C, or 40°C and 60°C, or 50°C and 60°C. In some embodiment of the invention, the mannanase polypeptide is active at 16°C or 32°C. In some embodiment of the invention, the mannanase polypeptide has mannanase activity in the presence of a protease. In some embodiments, the mannanase polypeptide retains activity in the presence of the bleaching agent. In some embodiments of the invention, the detergent composition having a mannanase polypeptide further comprises a bleaching agent. In some embodiment of the invention, the mannanase polypeptide has mannanase activity in the presence of a protease and an amylase. In some embodiments of the invention, the detergent composition having a mannanase polypeptide further comprises a protease and an amylase. [009] In some embodiments, the composition further comprises a surfactant. In some embodiments, the surfactant is selected from the group consisting of sodium dodecylbenzene sulfonate, sodium hydrogenated cocoate, sodium laureth sulfate, C12-14 pareth-7, C12-15 pareth-7, sodium C12-15 pareth sulfate, C14-15 pareth-4, and combinations thereof. In some embodiments, the surfactant is an ionic surfactant. In some embodiments, the ionic surfactant is selected from the group consisting of an anionic surfactant, a cationic surfactant, a zwitterionic surfactant, and a combination thereof. [0010] In some embodiments, the composition further comprises an enzyme selected from the group consisting proteases, proteases, peroxidases, cellulases, beta-glucanases, hemicellulases, lipases, acyl transferases, phospholipases, esterases, laccases, catalases, aryl esterases, amylases, alpha-amylases, glucoamylases, cutinases, pectinases, pectate lyases, keratinases, reductases, oxidases, phenoloxidases, lipoxygenases, ligninases, carrageenases, pullulanases, tannases, arabinosidases, hyaluronidases, chondroitinases, xyloglucanases, xylanases, pectin acetyl esterases, polygalacturonases, rhamnogalacturonases, other endo- -mannanases, exo- -mannanases, pectin methylesterases, cellobiohydrolases, transglutaminases, and combinations thereof.