ProGlyProt ID BC105 Organism Information Organism Name Bacillus lentus Domain Bacteria Classification Family: Bacillaceae Order: Bacillales Class: Bacilli (or Firmibacteria) Division or phylum: "Firmicutes" Taxonomic ID (NCBI) 1467 Genome Sequence (s) EMBL Gene Information Gene Name NCBI Gene ID Protein Information Protein Name Subtilisin (SBL)- Cys mutant UniProtKB/SwissProt ID  P29600 NCBI RefSeq EMBL-CDS UniProtKB Sequence >sp|P29600|SUBS_BACLE Subtilisin Savinase OS=Bacillus lentus PE=1 SV=1 AQSVPWGISRVQAPAAHNRGLTGSGVKVAVLDTGISTHPDLNIRGGASFVPGEPSTQDGN GHGTHVAGTIAALNNSIGVLGVAPSAELYAVKVLGASGSGSVSSIAQGLEWAGNNGMHVA NLSLGSPSPSATLEQAVNSATSRGVLVVAASGNSGAGSISYPARYANAMAVGATDQNNNR ASFSQYGAGLDIVAPGVNVQSTYPGSTYASLNGTSMATPHVAGAAALVKQKNPSWSNVQI RNHLKNTATSLGSTNLYGSGLVNAEAATR Sequence length 269 AA Subcellular Location Secreted Function Subtilisin is an extracellular alkaline serine protease. EC = 3.4.21.62 Protein Structure PDB ID 1JEA Glycosylation Status Glycosylation Type S (Cys) linked Experimentally Validated Glycosite(s) in Full Length Protein N62C, S156C, S166C and L217C (Residues that are shown glycosylated in vitro upon mutation from, N, S and L to Cys respectively) Experimentally Validated Glycosite(s ) in Mature Protein N62C, S156C, S166C and L217C (Residues that are shown glycosylated in vitro upon mutation from, N, S and L to Cys respectively) Glycosite(s) Annotated Protein Sequence >1JEA:A|PDBID|CHAIN|SEQUENCE AQSVPWGISRVQAPAAHNRGLTGSGVKVAVLDTGISTHPDLNIRGGASFVPGEPSTQDGNGHGTHVAGTIAALNNSIGVL GVAPSAELYAVKVLGASGSGSVSSIAQGLEWAGNNGMHVANLSLGSPSPSATLEQAVNSATSRGVLVVAASGNSGAGSIS YPARYANAMAVGATDQNNNRASFSQYGAGLDIVAPGVNVQSTYPGSTYASLNGTSMATPHVAGAAALVKQKNPSWSNVQI RNHLKNTATSLGSTNLYGSGLVNAEAATR This sequence was mutated at four residues to cysteines which were then glycosylated: >1JEA:A|PDBID|CHAIN|SEQUENCE AQSVPWGISRVQAPAAHNRGLTGSGVKVAVLDTGISTHPDLNIRGGASFVPGEPSTQDGC*(62)GHGTHVAGTIAALNNSIGVL GVAPSAELYAVKVLGASGSGSVSSIAQGLEWAGNNGMHVANLSLGSPSPSATLEQAVNSATSRGVLVVAASGNC*(156)GAGSIC*(166) YPARYANAMAVGATDQNNNRASFSQYGAGLDIVAPGVNVQSTYPGSTYASC*(217)NGTSMATPHVAGAAALVKQKNPSWSNVQI RNHLKNTATSLGSTNLYGSGLVNAEAATR Sequence Around Glycosites (21 AA) VPGEPSTQDGCGHGTHVAGTI GVLVVAASGNCGAGSISYPAR ASGNSGAGSICYPARYANAMA STYPGSTYASCNGTSMATPHV Glycosite Sequence Logo seqlogo Glycosite Sequence Logo Technique(s) used for Glycosylation Detection In vitro chemical glycosylation Technique(s) used for Glycosylated Residue(s) Detection Not applicable Protein Glycosylation- Implication In vitro engineered glycosylations of native subtilisin has been shown to affect structure as well as enzymatic activity. Glycan Information Glycan Annotation L217C-S-β-Glc(Ac)2, L217C-S-β-Glc(Ac)3, N62C-S-β-Glc(Ac)4, S156C-S-β-Glc(Ac)4, S16C-S-β-Glc(Ac)4. Technique(s) used for Glycan Identification Acetylated glycans were predetermined and synthesized in vitro Protein Glycosylation linked (PGL) gene(s) OST Gene Name OST NCBI Gene ID OST GenBank Gene Sequence OST Protein Name OST UniProtKB/ SwissProt ID OST NCBI RefSeq OST EMBL-CDS OST UniProtKB Sequence OST EC Number (BRENDA) OST Genome Context Characterized Accessory Gene(s) PGL Additional Links CAZy Literatures Reference(s) 1) Davis, B.G., Lloyd, R.C. and Jones, J.B. (2000) Controlled site-selective protein glycosylation for precise glycan structure-catalytic activity relationships. Bioorg Med Chem, 8, 1527-1535. [PubMed: 10976501] 2) Lloyd, R.C. Davis, B.G. and Jones, J.B. (2000) Site-selective glycosylation of subtilisin Bacillus lentus causes dramatic increases in esterase activity. Bioorg Med Chem, 8, 1537-44. [PubMed: 10976502] 3) Graycar, T., Knapp, M., Ganshaw, G., Dauberman, J. and Bott, R. (1999) Additional Comments Engineered glycoprotein. Site-selective cysteine (modified mutant) glycosylations of subtilisin, a Bacillus lentus (SBL) protein has been shown to affect structure as well as enzymatic activity post in vitro glycosylation. A positive correlation has also been derived between acetylation of glycan attached and the enzymatic activity and specificity against an esterase substrate succinyl-Ala-Ala-Pro-Phe-S-benzyl by glycosylated subtilisin. It is one of the first examples of preparations Year of Identification 2000 Year of Validation 2000