{"title":"设计量身定制的立体物质,提高固定化菲辛对小蛋白和大蛋白的特异性","authors":"","doi":"10.1016/j.jbiotec.2024.09.005","DOIUrl":null,"url":null,"abstract":"<div><p>The development of strategies that can permit to adjust the size specificity of immobilized proteases by the generation of steric hindrances may enlarge its applicability. Using as a model ficin immobilized on glyoxyl agarose, two strategies were assayed to generate tailor made steric hindrances. First, ficin has been coimmobilized on supports coated with large proteins (hemoglobin or bovine serum albumin (BSA)). While coimmobilization of ficin with BSA presented no effect on the activity versus any of the assayed substrates, coimmobilization with hemoglobin permitted to improve the immobilized ficin specificity for casein versus hemoglobin, but still significant activity versus hemoglobin remained. Second, aldehyde-dextran has been employed to modify the immobilized ficin, trying to generate steric hindrances to avoid the entry of large proteins (hemoglobin) while enabling the entry of small ones (casein). This also increased the size specificity of ficin, but still did not suppress the activity versus hemoglobin. The combination of both strategies and the use of 37ºC during the proteolysis enabled to almost fully nullify the hydrolytic activity versus hemoglobin while preserving a high percentage of the activity versus casein. The modifications improved enzyme stability and the biocatalyst could be reused for 5 cycles without alteration of its properties.</p></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":null,"pages":null},"PeriodicalIF":4.1000,"publicationDate":"2024-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S0168165624002475/pdfft?md5=701784417039f9dca9ae5e5f9b351c9f&pid=1-s2.0-S0168165624002475-main.pdf","citationCount":"0","resultStr":"{\"title\":\"Designing tailor-made steric matters to improve the immobilized ficin specificity for small versus large proteins\",\"authors\":\"\",\"doi\":\"10.1016/j.jbiotec.2024.09.005\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>The development of strategies that can permit to adjust the size specificity of immobilized proteases by the generation of steric hindrances may enlarge its applicability. Using as a model ficin immobilized on glyoxyl agarose, two strategies were assayed to generate tailor made steric hindrances. First, ficin has been coimmobilized on supports coated with large proteins (hemoglobin or bovine serum albumin (BSA)). While coimmobilization of ficin with BSA presented no effect on the activity versus any of the assayed substrates, coimmobilization with hemoglobin permitted to improve the immobilized ficin specificity for casein versus hemoglobin, but still significant activity versus hemoglobin remained. Second, aldehyde-dextran has been employed to modify the immobilized ficin, trying to generate steric hindrances to avoid the entry of large proteins (hemoglobin) while enabling the entry of small ones (casein). This also increased the size specificity of ficin, but still did not suppress the activity versus hemoglobin. The combination of both strategies and the use of 37ºC during the proteolysis enabled to almost fully nullify the hydrolytic activity versus hemoglobin while preserving a high percentage of the activity versus casein. The modifications improved enzyme stability and the biocatalyst could be reused for 5 cycles without alteration of its properties.</p></div>\",\"PeriodicalId\":15153,\"journal\":{\"name\":\"Journal of biotechnology\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":4.1000,\"publicationDate\":\"2024-09-10\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.sciencedirect.com/science/article/pii/S0168165624002475/pdfft?md5=701784417039f9dca9ae5e5f9b351c9f&pid=1-s2.0-S0168165624002475-main.pdf\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Journal of biotechnology\",\"FirstCategoryId\":\"5\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0168165624002475\",\"RegionNum\":2,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"BIOTECHNOLOGY & APPLIED MICROBIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of biotechnology","FirstCategoryId":"5","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0168165624002475","RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
Designing tailor-made steric matters to improve the immobilized ficin specificity for small versus large proteins
The development of strategies that can permit to adjust the size specificity of immobilized proteases by the generation of steric hindrances may enlarge its applicability. Using as a model ficin immobilized on glyoxyl agarose, two strategies were assayed to generate tailor made steric hindrances. First, ficin has been coimmobilized on supports coated with large proteins (hemoglobin or bovine serum albumin (BSA)). While coimmobilization of ficin with BSA presented no effect on the activity versus any of the assayed substrates, coimmobilization with hemoglobin permitted to improve the immobilized ficin specificity for casein versus hemoglobin, but still significant activity versus hemoglobin remained. Second, aldehyde-dextran has been employed to modify the immobilized ficin, trying to generate steric hindrances to avoid the entry of large proteins (hemoglobin) while enabling the entry of small ones (casein). This also increased the size specificity of ficin, but still did not suppress the activity versus hemoglobin. The combination of both strategies and the use of 37ºC during the proteolysis enabled to almost fully nullify the hydrolytic activity versus hemoglobin while preserving a high percentage of the activity versus casein. The modifications improved enzyme stability and the biocatalyst could be reused for 5 cycles without alteration of its properties.
期刊介绍:
The Journal of Biotechnology has an open access mirror journal, the Journal of Biotechnology: X, sharing the same aims and scope, editorial team, submission system and rigorous peer review.
The Journal provides a medium for the rapid publication of both full-length articles and short communications on novel and innovative aspects of biotechnology. The Journal will accept papers ranging from genetic or molecular biological positions to those covering biochemical, chemical or bioprocess engineering aspects as well as computer application of new software concepts, provided that in each case the material is directly relevant to biotechnological systems. Papers presenting information of a multidisciplinary nature that would not be suitable for publication in a journal devoted to a single discipline, are particularly welcome.