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ProGlyProt ID
BC153
Organism Information
Organism Name
Mycobacterium tuberculosis H37Rv
Domain
Bacteria
Classification
Family: Mycobacteriaceae
Suborder: Corynebacterineae
Order: Actinomycetales
Subclass: Actinobacteridae
Class: Actinobacteria
Division or phylum: "Actinobacteria"
Taxonomic ID (NCBI)
 
 
Genome Sequence(s)
GenBank
EMBL
Organism Additional Information
It is the causative agent of human tuberculosis. The pathogenesis is influenced by its lipoglycans and glycolipids (having a wide range of immunomodulatory activities), and a variety of its virulence factors and antigens.
 
 
Gene Information
Gene Name
Apa (Rv1860) or modD
NCBI Gene ID
GenBank Gene Sequence
 
 
Protein Information
Protein Name
Alanine and proline-rich secreted protein Apa (50/55-kDa or 45 kDa MPT 32)
UniProtKB/SwissProt ID
NCBI RefSeq
EMBL-CDS
UniProtKB Sequence
Sequence length
325 AA
Subcellular Location
Secreted
Function
Immunodominant antigen. Elicits potent DTH (delayed-type hypersensitivity) responses in living-BCG-immunized guinea pigs..Stimulation of peripheral blood mononuclear cells from PPD positive individuals with Apa induces a Th1 type lymphoproliferative response with expansion of both CD4+ and CD8+ cells and increased IFN-γ production.
 
 
Glycosylation Status
Glycosylation Type
O (Thr) linked
Experimentally Validated Glycosite(s) in Full Length Protein
(Signal peptide: 1-39) T49, T57, T66, T316
Experimentally Validated Glycosite(s ) in Mature Protein
T10, T18, T27, T277
Glycosite(s) Annotated Protein Sequence
>sp|Q50906|APA_MYCTU Alanine and proline-rich secreted protein apa OS=Mycobacter
ium tuberculosis GN=apa PE=1 SV=1 MHQVDPNLTRRKGRLAALAIAAMASASLVTVAVPATANADP
EPAPPVPT*(49)TAASPPST*(57)AAA PPAPAT*(66)PVAPPPPAAANTPNAQPGDPNAAPPPAD
PNAPPPPVIAPNAPQPVRIDNPVGGF SFALPAGWVESDAAHFDYGSALLSKTTGDPPFPGQPPPVANDTRIVLG
RLDQKLYASAEA TDSKAAARLGSDMGEFYMPYPGTRINQETVSLDANGVSGSASYYEVKFSDPSKPNGQIWT
GVIGSPAANAPDAGPPQRWFVVWLGTANNPVDKGAAKALAESIRPLVAPPPAPAPAPAEP APAPAPAGEVAPTP
TT*(316)PTPQRTLPA
Sequence Around Glycosites (21 AA)
ADPEPAPPVPTTAASPPSTAA
VPTTAASPPSTAAAPPAPATP
STAAAPPAPATPVAPPPPAAA
PAGEVAPTPTTPTPQRTLPA
Glycosite Sequence Logo
Glycosite Sequence Logo
Technique(s) used for Glycosylation Detection
Schiff's staining, peroxidase-conjugated concanavalin A (ConA)-binding
Technique(s) used for Glycosylated Residue(s) Detection
N-terminal amino acid sequencing coupled with fast atom bombardment-mass spectrometry (FAB-MS)
Protein Glycosylation- Implication
The presence of the mannose residues on the Apa protein was essential for the antigenicity of the molecules in T-cell-dependent immune responses in vitro (proliferation assay) and in vivo (delayed type hypersensitivity or DTH reaction). The deglycosylated antigen was 10-fold less active than native molecules in eliciting DTH. Mannosylation of antigen leads to selective targeting and subsequent greater presentation by dendritic cells.
 
 
Glycan Information
Glycan Annotation
Linkage: αMan-Thr.
34.6% sugar detected.
α-D-Manp(1→2)α-D-Manp (mannobiose), α-D-Manp (single mannose), α-D-Manp(1→2)α-D-Manp(1→2)α-D-Manp (mannotriose) are present. T10 and T18 are glycosylated with mannobiose, T27 with single mannose, and T277 with either of the three. The majority of the antigen species bear six, seven, or eight mannose residues (22, 24, and 17%, respectively), while others three, four, or five mannoses (5, 9, and 14%, respectively).
Technique(s) used for Glycan Identification
GC-MS of partially methylated alditol acetates obtained by trifluoro acetic acid (TFA) hydrolysis of permethylated oligoglycosyl alditols which are released by β-elimination of the glycopeptides.
 
 
Protein Glycosylation linked (PGL) gene(s)
OST Gene Name
pmt/rv1002c
OST NCBI Gene ID
OST GenBank Gene Sequence
OST Protein Name
Protein O mannosyltransferase
OST UniProtKB/ SwissProt ID
OST NCBI RefSeq
OST EMBL-CDS
OST UniProtKB Sequence
OST EC Number (BRENDA)
OST Genome Context
Characterized Accessory Gene(s)
Polyprenol-phosphate-mannose (PPM) synthase, Ppm1, is present. A second type of Ppm synthase (Rv3779 gene product) exclusive to slow-growing mycobacteria, is a membrane glycosyltransferase. It mannosylates polyprenyl-phosphates directly from GDP-mannose.
PGL Additional Links
 
 
Literature
Reference(s)
1) Scherman, H., Kaur, D., Pham, H., Skovierova, H., Jackson, M. and Brennan, P.J. (2009) Identification of a polyprenylphosphomannosyl synthase involved in the synthesis of mycobacterial mannosides. J Bacteriol, 191, 6769-6772. [PubMed: 19717608]
2) Lara, M., Servin-Gonzalez, L., Singh, M., Moreno, C., Cohen, I., Nimtz, M. and Espitia, C. (2004) Expression, secretion, and glycosylation of the 45- and 47-kDa glycoprotein of Mycobacterium tuberculosis in Streptomyces lividans. Appl Environ Microbiol, 70, 679-685. [PubMed: 14766542]
3) Cooper, H.N., Gurcha, S.S., Nigou, J., Brennan, P.J., Belisle, J.T., Besra, G.S. and Young, D. (2002) Characterization of mycobacterial protein glycosyltransferase activity using synthetic peptide acceptors in a cell-free assay. Glycobiology, 12, 427-434. [PubMed: 12122024]
4) Horn, C., Namane, A., Pescher, P., Riviere, M., Romain, F., Puzo, G., Barzu, O. and Marchal, G. (1999) Decreased capacity of recombinant 45/47-kDa molecules (Apa) of Mycobacterium tuberculosis to stimulate T lymphocyte responses related to changes in their mannosylation pattern. J Biol Chem, 274, 32023-32030. [PubMed: 10542234]
5) Romain, F., Horn, C., Pescher, P., Namane, A., Riviere, M., Puzo, G., Barzu, O. and Marchal, G. (1999) Deglycosylation of the 45/47-kilodalton antigen complex of Mycobacterium tuberculosis decreases its capacity to elicit in vivo or in vitro cellular immune responses. Infect Immun, 67, 5567-5572. [PubMed: 10531201]
6) Dobos, K.M., Khoo, K.H., Swiderek, K.M., Brennan, P.J. and Belisle, J.T. (1996) Definition of the full extent of glycosylation of the 45-kilodalton glycoprotein of Mycobacterium tuberculosis. J Bacteriol, 178, 2498-2506. [PubMed: 8626314]
7) Dobos, K.M., Swiderek, K., Khoo, K.H., Brennan, P.J. and Belisle, J.T. (1995) Evidence for glycosylation sites on the 45-kilodalton glycoprotein of Mycobacterium tuberculosis. Infect Immun, 63, 2846-2853. [PubMed: 7622204]
8) Espitia, C., Espinosa, R., Saavedra, R., Mancilla, R., Romain, F., Laqueyrerie, A. and Moreno, C. (1995) Antigenic and structural similarities between Mycobacterium tuberculosis 50- to 55-kilodalton and Mycobacterium bovis BCG 45- to 47-kilodalton antigens. Infect Immun, 63, 580-584. [PubMed: 7822025]
9) Espitia, C. and Mancilla, R. (1989) Identification, isolation and partial characterization of Mycobacterium tuberculosis glycoprotein antigens. Clin Exp Immunol, 77, 378-383. [PubMed: 2478323]
Additional Comments
Sec-mediated translocation influences the O-mannosylation. Ppm1 does not discriminate between polyprenol substrates with variable chain lengths and saturation of the isoprene units.
Glycosylation sites have been found to be located within proline-rich domains (or Thr-rich sequences) near the N-terminus and the C-terminus.
In a cell-free assay, the M. smegmatis mannosyltransferase activity in membrane and cell wall fraction has been shown to catalyze transfer of radiolabeled mannose from GDP-[14C]mannose to peptide acceptors. The acceptors consisted of Thr-rich sequences from M. tuberculosis 45 kDa antigen (Ref. no. 3).
The 45/47 kDa antigen (Apa) has also been glycosylated in Streptomyces lividans. The 45- and 47-kDa represent two glycoforms of the antigen (Ref. no. 2).
Year of Identification
1989
Year of Validation
1996
 
 
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