Random Mutagenesis Identifies Novel Genes Involved in the Secretion of Antimicrobial, Cell Wall-Lytic Enzymes by Lactococcus lactis

Article


Tan, Yu Pei, Giffard, Philip M., Barry, Daniel G., Huston, Wilhelmina M. and Turner, Mark S.. 2008. "Random Mutagenesis Identifies Novel Genes Involved in the Secretion of Antimicrobial, Cell Wall-Lytic Enzymes by Lactococcus lactis." Applied and Environmental Microbiology. 74 (24), pp. 7490-7496. https://doi.org/10.1128/AEM.00767-08
Article Title

Random Mutagenesis Identifies Novel Genes Involved in the Secretion of Antimicrobial, Cell Wall-Lytic Enzymes by Lactococcus lactis

ERA Journal ID2456
Article CategoryArticle
AuthorsTan, Yu Pei, Giffard, Philip M., Barry, Daniel G., Huston, Wilhelmina M. and Turner, Mark S.
Journal TitleApplied and Environmental Microbiology
Journal Citation74 (24), pp. 7490-7496
Number of Pages7
YearDec 2008
PublisherAmerican Society for Microbiology
Place of PublicationUnited States
ISSN0099-2240
1098-5336
Digital Object Identifier (DOI)https://doi.org/10.1128/AEM.00767-08
Web Address (URL)https://journals.asm.org/doi/10.1128/AEM.00767-08
Abstract

Lactococcus lactis is a gram-positive bacterium that is widely used in the food industry and is therefore desirable as a candidate for the production and secretion of recombinant proteins. Previously, we generated a L. lactis strain that expressed and secreted the antimicrobial cell wall-lytic enzyme lysostaphin. To identify lactococcal gene products that affect the production of lysostaphin, we isolated and characterized mutants generated by random transposon mutagenesis that had altered lysostaphin activity. Out of 35,000 mutants screened, only one with no lysostaphin activity was identified, and it was found to contain an insertion in the lysostaphin expression cassette. Ten mutants with higher lysostaphin activity contained insertions in only four different genes, which encode an uncharacterized putative transmembrane protein (llmg_0609) (three mutants), an enzyme catalyzing the first step in peptidoglycan biosynthesis (murA2) (five mutants), a putative regulator of peptidoglycan modification (trmA) (one mutant), and an uncharacterized enzyme possibly involved in ubiquinone biosynthesis (llmg_2148) (one mutant). These mutants were found to secrete larger amounts of lysostaphin than the control strain (MG1363[lss]), and the greatest increase in secretion was 9.8- to 16.1-fold, for the llmg_0609 mutants. The lysostaphinoversecreting llmg_0609, murA2, and trmA mutants were also found to secrete larger amounts of another cell wall-lytic enzyme (the Listeria monocytogenes bacteriophage endolysin Ply511) than the control strain, indicating that the phenotype is not limited to lysostaphin.

KeywordsBiochemical engineering; Biochemistry; Biosynthesis; Enzymes; Genes; Mutagenesis; Proteins
Contains Sensitive ContentDoes not contain sensitive content
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Byline AffiliationsQueensland University of Technology
Menzies School of Health Research, Australia
University of Queensland
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Dann, E. K., Cooke, A. W., Forsberg, L. I., Pegg, K. G., Tan, Y. P. and Shivas, R. G.. 2012. "Pathogenicity studies in avocado with three nectriaceous fungi, Calonectria ilicicola, Gliocladiopsis sp. and Ilyonectria liriodendri." Plant Pathology. 61 (5), pp. 896-902. https://doi.org/10.1111/j.1365-3059.2011.02579.x
Fungal planet description sheets: 128-153
Crous, P. W., Shivas, R. G., Wingfield, M. J., Summerell, B. A., Rossman, A. Y., Alves, J. L., Adams, G. C., Barreto, R. W., Bell, A., Coutinho, M. L., Lombard, L., Quaedvlieg, W., Videira, S. I R, Groenewald, J. Z., Tan, Y. P., Mahoney, D. P., Macedo, F., Flory, S. L., Gates, G., ..., Walker, D. M.. 2012. "Fungal planet description sheets: 128-153." Persoonia. 29, pp. 146-201. https://doi.org/10.3767/003158512X661589
Fungal planet description sheets: 214-280
Crous, P. W., Shivas, R. G., Quaedvlieg, W., Lombard, L., Groenewald, J. Z., van der Bank, M., Zhang, Y., Summerell, B. A., Guarro, J., Wingfield, M. J., Tan, Y. P., Cano-Lira, J. F., García, D., Marin-Felix, Y., Alvarado, P., Gené, J., Hernández-Restrepo, M., Marin-Felix, Y., Stchigel, A. M., ..., Wong, P. T. W.. 2014. "Fungal planet description sheets: 214-280." Persoonia. 32, pp. 184-306. https://doi.org/10.3767/003158514X682395
Fungal planet description sheets: 371–399
Crous, P. W., Wingfield, M. J., Le Roux, J. J., Richardson, D. M., Strasberg, D., Shivas, R. G., Alvarado, P., Edwards, J., Moreno, G., Sharma, R., Sonawane, M. S., Tan, Y. P., Altes, A., Barasubiye, T., Barnes, C. W., Blanchette, R. A., Boertmann, D., Bogo, A., Carlavilla, J. R., ..., Groenewald, J. Z.. 2015. "Fungal planet description sheets: 371–399." Persoonia. 35, pp. 264-327. https://doi.org/10.3767/003158515X690269
Fungal planet description sheets: 469-557
Crous, P. W., Wingfield, M. J., Burgess, T. I., Hardy, G. E. St. J., Crane, C., Barrett, S., Cano-Lira, J. F., Le Roux, J. J., Thangavel, R., Guarro, J., Stchigel, A. M., Martin, M. P., Alfredo, D. S., Barber, P. A., Barreto, R. W., Baseia, I. G., Cano-Canals, J., Cheewangkoon, R., Ferreira, R. J., ..., Groenewald, J. Z.. 2016. "Fungal planet description sheets: 469-557." Persoonia. 37, pp. 218-403. https://doi.org/10.3767/003158516X694499
Colletotrichum siamense, C. theobromicola and C. queenslandicum, from several plant species and the identification of C. asianum in the Northern Territory, Australia
James, R. S., Ray, J., Tan, Y. P. and Shivas, R. G.. 2014. "Colletotrichum siamense, C. theobromicola and C. queenslandicum, from several plant species and the identification of C. asianum in the Northern Territory, Australia." Australasian Plant Disease Notes. 9 (1), pp. 138-138. https://doi.org/10.1007/s13314-014-0138-x
Elucidation of the taxonomy and pathological status of Pyricularia associated with banana blast in Australia
Male, Mike F., Tan, Yu Pei, Vawdrey, Lynton L. and Shivas, Roger G.. 2011. "Elucidation of the taxonomy and pathological status of Pyricularia associated with banana blast in Australia." Australasian Plant Disease Notes. 6 (1), pp. 22-25. https://doi.org/10.1007/s13314-011-0008-8
First report of shoot blight of persimmon caused by Diaporthe neotheicola in Australia
Golzar, H., Wang, C., Tan, Y. P. and Shivas, R. G.. 2012. "First report of shoot blight of persimmon caused by Diaporthe neotheicola in Australia." Australasian Plant Disease Notes. 7 (1), pp. 115-117. https://doi.org/10.1007/s13314-012-0061-y
First report of Phoma multirostrata in Australia
Golzar, H., Lanoiselet, V., Wang, C., Tan, Y. P. and Shivas, R. G.. 2015. "First report of Phoma multirostrata in Australia." Australasian Plant Disease Notes. 10 (1), pp. 8-9. https://doi.org/10.1007/s13314-015-0161-6
A new host genus and species (Afzelia xylocarpa) for Phakopsora pachyrhizi found in Thailand
Seemadua, Suneerat, Bhasabutra, Tharntip, Beasley, Dean R., Tan, Yu Pei and Shivas, Roger G.. 2012. "A new host genus and species (Afzelia xylocarpa) for Phakopsora pachyrhizi found in Thailand." Australasian Plant Disease Notes. 7 (1), pp. 125-126. https://doi.org/10.1007/s13314-012-0064-8
Recovery, pathogenicity and molecular sequencing of Calonectria ilicicola which causes collar rot on Carica papaya in Australia
Male, M. F., Tan, Y. P., Vawdrey, L. L. and Shivas, R. G.. 2012. "Recovery, pathogenicity and molecular sequencing of Calonectria ilicicola which causes collar rot on Carica papaya in Australia." Australasian Plant Disease Notes. 7 (1), pp. 137-138. https://doi.org/10.1007/s13314-012-0068-4
Review of Fusarium species isolated in association with mango malformation in Australia
Liew, E. C. Y., Laurence, M. H., Pearce, C. A., Shivas, R. G., Johnson, G. I., Tan, Y. P., Edwards, J., Perry, S., Cooke, A. W. and Summerell, B. A.. 2016. "Review of Fusarium species isolated in association with mango malformation in Australia." Australasian Plant Pathology. 45 (6), pp. 547-559. https://doi.org/10.1007/s13313-016-0454-z
Detection of Fusarium oxysporum f. sp. cubense tropical race 4 strain in northern Queensland
O'Neill, W. T., Pattemore, J. A., Perry, S., Beasley, D. R., Tan, Y. P., Smyth, A. L., Goosem, C. H., Thomson, K. M., Hobbs, R. L., Shivas, R. G., Henderson, J., O'Dwyer, C., Grice, K. R. E., Trevorrow, P., Pathania, N. and Vawdrey, L. L.. 2016. "Detection of Fusarium oxysporum f. sp. cubense tropical race 4 strain in northern Queensland." Australasian Plant Disease Notes. 11 (33), pp. 33-35. https://doi.org/10.1007/s13314-016-0218-1
Green and brown bridges between weeds and crops reveal novel Diaporthe species in Australia
Thompson, S. M., Tan, Y. P., Shivas, R. G., Neate, S. M., Morin, L., Bissett, A. and Aitken, E. A. B.. 2015. "Green and brown bridges between weeds and crops reveal novel Diaporthe species in Australia." Persoonia. 35, pp. 39-49. https://doi.org/10.3767/003158515X687506
Fungal Planet description sheets: 281-319
Crous, P., Wingfield, M., Schumacher, R., Summerell, B., Giraldo, A., Gene, J., Guarro, J., Wanasinghe, D., Hyde, K., Camporesi, E., Jones, E., Thambugala, K., Malysheva, E., Malysheva, V., Acharya, K., Alvarez, J., Alvarado, P., Assefa, A., Barnes, C. W., ..., Groenewald, J.. 2014. "Fungal Planet description sheets: 281-319." Persoonia. 33, pp. 212-292. https://doi.org/10.3767/003158514X685680
Fusarium thapsinum is the dominant species associated with sorghum stalk rot in Queensland and northern New South Wales
Kelly, L., Tan, Y. P., Ryley, M. and Aitken, E.. 2011. "Fusarium thapsinum is the dominant species associated with sorghum stalk rot in Queensland and northern New South Wales." Tran-Nguyen, Lucy (ed.) 4th Asian Conference on Plant Pathology and the 18th Biennial Australasian Plant Pathology Society Conference (ACPP/APPS 2011): New Frontiers in Plant Pathology for Asia and Oceania. Darwin, Australia 26 - 29 Apr 2011 Toowoomba, Australia.
Stem cankers on sunflower (Helianthus annuus) in Australia reveal a complex of pathogenic Diaporthe (Phomopsis) species
Thompson, S. M., Tan, Y. P., Young, A. J., Neate, S. M., Aitken, E. A. B. and Shivas, R. G.. 2011. "Stem cankers on sunflower (Helianthus annuus) in Australia reveal a complex of pathogenic Diaporthe (Phomopsis) species." Persoonia. 27, pp. 80-89. https://doi.org/10.3767/003158511X617110
Phyllosticta spp. on cultivated Citrus in Australia
Miles, A. K., Tan, Y. P., Tan, M. K., Donovan, N. J., Ghalayini, A. and Drenth, A.. 2013. "Phyllosticta spp. on cultivated Citrus in Australia ." Australasian Plant Pathology. 42 (4), pp. 461-467. https://doi.org/10.1007/s13313-013-0208-0
A case for re-inventory of Australia's plant pathogens
Hyde, K. D., Chomnunti, P., Crous, P. W., Groenewald, J. Z., Damm, U., Ko, T. W. Ko, Shivas, R. G., Summerell, B. A. and Tan, Y. P.. 2010. "A case for re-inventory of Australia's plant pathogens." Persoonia. 25, pp. 50-60. https://doi.org/10.3767/003158510X548668
Austropleospora osteospermi gen. et sp. nov. and its host specificity and distribution on Chrysanthemoides monilifera ssp. rotundata in Australia
Morin, Louise, Shivas, R. G., Piper, M. C. and Tan, Y. P.. 2010. "Austropleospora osteospermi gen. et sp. nov. and its host specificity and distribution on Chrysanthemoides monilifera ssp. rotundata in Australia ." Fungal Diversity. 40 (1), pp. 65-74. https://doi.org/10.1007/s13225-009-0007-7
Inactivation of an Iron Transporter in Lactococcus lactis Results in Resistance to Tellurite and Oxidative Stress
Turner, Mark S., Tan, Yu Pei and Giffard, Philip M.. 2007. "Inactivation of an Iron Transporter in Lactococcus lactis Results in Resistance to Tellurite and Oxidative Stress." Applied and Environmental Microbiology. 73 (19), pp. 6144-6149. https://doi.org/10.1128/AEM.00413-07