Proteolytic profiles of two isoforms of human AMBN expressed in E. coli by MMP-20 and KLK-4 proteases
Ameloblastin is a protein in biomineralization of tooth enamel. However recent results indicate that this is probably not its only role in an organism. Enamel matrix formation represents a complex process enabled via specific crosslinking of two proteins – the most abundant amelogenin and the amelob...
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Elsevier
2024-01-01
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2405844024005954 |
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| author | Vetyskova Veronika Hubalek Martin Sulc Josef Prochazka Jan Vondrasek Jiri Kristyna Vydra Bousova |
| author_facet | Vetyskova Veronika Hubalek Martin Sulc Josef Prochazka Jan Vondrasek Jiri Kristyna Vydra Bousova |
| author_sort | Vetyskova Veronika |
| collection | DOAJ |
| description | Ameloblastin is a protein in biomineralization of tooth enamel. However recent results indicate that this is probably not its only role in an organism. Enamel matrix formation represents a complex process enabled via specific crosslinking of two proteins – the most abundant amelogenin and the ameloblastin (AMBN). The human AMBN (hAMBN) gene possesses 13 protein coding exons with alternatively spliced transcripts and the longest isoform about 447 amino acid residues. It has been described that AMBN molecules in vitro assemble into oligomers via a sequence encoded by exon 5. Enamel is formed by the processing of enamel proteins by two specific proteases - enamelysin (MMP-20) and kallikrein 4 (KLK-4). The scaffold made of AMEL and non-amelogenin proteins is cleaved and removed from the developed tooth enamel. The hAMBN is expressed in two isoforms (ISO I and II), which could lead to their different utilization determined by distinct proteolytic profiles. In this study, we compared proteolytic profiles of both isoforms of hAMBN expressed in E. coli after proteolysis by MMP-20, KLK-4, and their 1:2 mixture. Proteolysis products were analysed and cleavage sites were identified by mass spectrometry. The proteolytic profiles of two AMBN isoforms showed different results, although we have to determine that the analysed AMBN was not post-translationally modified as expressed in prokaryotic cells. These results may lead to the suggestion of potentially divergent roles of AMBN isoforms cleavage products in various cell signalling pathways such as calcium buffering or signalling cascades. |
| first_indexed | 2024-03-08T06:55:18Z |
| format | Article |
| id | doaj.art-bfdd9bbe7ecc46f49422ffbb3aab2c47 |
| institution | Directory Open Access Journal |
| issn | 2405-8440 |
| language | English |
| last_indexed | 2024-03-08T06:55:18Z |
| publishDate | 2024-01-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Heliyon |
| spelling | doaj.art-bfdd9bbe7ecc46f49422ffbb3aab2c472024-02-03T06:37:52ZengElsevierHeliyon2405-84402024-01-01102e24564Proteolytic profiles of two isoforms of human AMBN expressed in E. coli by MMP-20 and KLK-4 proteasesVetyskova Veronika0Hubalek Martin1Sulc Josef2Prochazka Jan3Vondrasek Jiri4Kristyna Vydra Bousova5Institute of Organic Chemistry and Biochemistry of the Czech Academy of Sciences, Flemingovo namesti 2, 16000, Prague, Czech RepublicInstitute of Organic Chemistry and Biochemistry of the Czech Academy of Sciences, Flemingovo namesti 2, 16000, Prague, Czech RepublicInstitute of Organic Chemistry and Biochemistry of the Czech Academy of Sciences, Flemingovo namesti 2, 16000, Prague, Czech Republic; Department of Physical and Macromolecular Chemistry, Faculty of Natural Sciences, Charles University, Hlavova 8, 128 00 Prague 2, Czech RepublicInstitute of Molecular Genetics of the Czech Academy of Sciences, Videnska 5, 14000, Prague, Czech RepublicInstitute of Organic Chemistry and Biochemistry of the Czech Academy of Sciences, Flemingovo namesti 2, 16000, Prague, Czech Republic; Corresponding author.Institute of Organic Chemistry and Biochemistry of the Czech Academy of Sciences, Flemingovo namesti 2, 16000, Prague, Czech Republic; Corresponding author.Ameloblastin is a protein in biomineralization of tooth enamel. However recent results indicate that this is probably not its only role in an organism. Enamel matrix formation represents a complex process enabled via specific crosslinking of two proteins – the most abundant amelogenin and the ameloblastin (AMBN). The human AMBN (hAMBN) gene possesses 13 protein coding exons with alternatively spliced transcripts and the longest isoform about 447 amino acid residues. It has been described that AMBN molecules in vitro assemble into oligomers via a sequence encoded by exon 5. Enamel is formed by the processing of enamel proteins by two specific proteases - enamelysin (MMP-20) and kallikrein 4 (KLK-4). The scaffold made of AMEL and non-amelogenin proteins is cleaved and removed from the developed tooth enamel. The hAMBN is expressed in two isoforms (ISO I and II), which could lead to their different utilization determined by distinct proteolytic profiles. In this study, we compared proteolytic profiles of both isoforms of hAMBN expressed in E. coli after proteolysis by MMP-20, KLK-4, and their 1:2 mixture. Proteolysis products were analysed and cleavage sites were identified by mass spectrometry. The proteolytic profiles of two AMBN isoforms showed different results, although we have to determine that the analysed AMBN was not post-translationally modified as expressed in prokaryotic cells. These results may lead to the suggestion of potentially divergent roles of AMBN isoforms cleavage products in various cell signalling pathways such as calcium buffering or signalling cascades.http://www.sciencedirect.com/science/article/pii/S2405844024005954AmeloblastinProteolytic analysisMMP-20KLK-4Enzymatic cleavage products |
| spellingShingle | Vetyskova Veronika Hubalek Martin Sulc Josef Prochazka Jan Vondrasek Jiri Kristyna Vydra Bousova Proteolytic profiles of two isoforms of human AMBN expressed in E. coli by MMP-20 and KLK-4 proteases Heliyon Ameloblastin Proteolytic analysis MMP-20 KLK-4 Enzymatic cleavage products |
| title | Proteolytic profiles of two isoforms of human AMBN expressed in E. coli by MMP-20 and KLK-4 proteases |
| title_full | Proteolytic profiles of two isoforms of human AMBN expressed in E. coli by MMP-20 and KLK-4 proteases |
| title_fullStr | Proteolytic profiles of two isoforms of human AMBN expressed in E. coli by MMP-20 and KLK-4 proteases |
| title_full_unstemmed | Proteolytic profiles of two isoforms of human AMBN expressed in E. coli by MMP-20 and KLK-4 proteases |
| title_short | Proteolytic profiles of two isoforms of human AMBN expressed in E. coli by MMP-20 and KLK-4 proteases |
| title_sort | proteolytic profiles of two isoforms of human ambn expressed in e coli by mmp 20 and klk 4 proteases |
| topic | Ameloblastin Proteolytic analysis MMP-20 KLK-4 Enzymatic cleavage products |
| url | http://www.sciencedirect.com/science/article/pii/S2405844024005954 |
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