MINT MINT, Dpt of Biology, University of Rome Tor Vergata MINT http://mint.bio.uniroma2.it/mint url>http://mint.bio.uniroma2.it/mint/ https://mint.bio.uniroma2.it MINT-[0-9]+ https://mint.bio.uniroma2.it/index.php/results-interactions/?id=${ac} The Molecular INTeraction database (MINT) is a relational database designed to store interactions between biological molecules. http://mint.bio.uniroma2.it Functional characterization of DnaB helicase and its modulation by single-stranded DNA-binding protein in Mycobacterium tuberculosis Functional characterization of DnaB helicase and its modulation by single-stranded DNA-binding protein in Mycobacterium tuberculosis 2014 imex curation he.zhengguo@hotmail.com; hezhengguo@mail.hzau.edu.cn 10-Mar-2014: Contacted by IntAct-Help. Only protein-protein interactions ecoli Escherichia coli (strain K12) 2 hybrid two hybrid 2H 2h Gal4 transcription regeneration two-hybrid yeast two hybrid 2-hybrid classical two hybrid classical two hybrid 2H two-hybrid 2-hybrid Gal4 transcription regeneration 2h yeast two hybrid Y2H Y-2H 2 hybrid predetermined predetermined participant predetermined he.zhengguo@hotmail.com; hezhengguo@mail.hzau.edu.cn Accepted 2014-JAN-13 by LUANA 2014 imex curation Functional characterization of DnaB helicase and its modulation by single-stranded DNA-binding protein in Mycobacterium tuberculosis Functional characterization of DnaB helicase and its modulation by single-stranded DNA-binding protein in Mycobacterium tuberculosis 2014 imex curation he.zhengguo@hotmail.com; hezhengguo@mail.hzau.edu.cn 10-Mar-2014: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro spr surface plasmon resonance Optical biosensor BIAcore(r) BIAcore(r) Optical biosensor spr predetermined predetermined participant predetermined he.zhengguo@hotmail.com; hezhengguo@mail.hzau.edu.cn Accepted 2014-JAN-13 by LUANA 2014 imex curation Functional characterization of DnaB helicase and its modulation by single-stranded DNA-binding protein in Mycobacterium tuberculosis Functional characterization of DnaB helicase and its modulation by single-stranded DNA-binding protein in Mycobacterium tuberculosis 2014 imex curation he.zhengguo@hotmail.com; hezhengguo@mail.hzau.edu.cn 10-Mar-2014: Contacted by IntAct-Help. Only protein-protein interactions mycbp mycbp crosslink cross-linking study crosslink western blot western blot Immuno blot Immuno blot he.zhengguo@hotmail.com; hezhengguo@mail.hzau.edu.cn Accepted 2014-JAN-13 by LUANA 2014 imex curation dnab_myctu Replicative DNA helicase dnaB MTCY21D4.21 Rv0058 protein protein myctu Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv) MAVVDDLAPGMDSSPPSEDYGRQPPQDLAAEQSVLGGMLLSKDAIADVLERLRPGDFYRPAHQNVYDAILDLYGRGEPADAVTVAAELDRRGLLRRIGGAPYLHTLISTVPTAANAGYYASIVAEKALLRRLVEAGTRVVQYGYAGAEGADVAEVVDRAQAEIYDVADRRLSEDFVALEDLLQPTMDEIDAIASSGGLARGVATGFTELDEVTNGLHPGQMVIVAARPGVGKSTLGLDFMRSCSIRHRMASVIFSLEMSKSEIVMRLLSAEAKIKLSDMRSGRMSDDDWTRLARRMSEISEAPLFIDDSPNLTMMEIRAKARRLRQKANLKLIVVDYLQLMTSGKKYESRQVEVSEFSRHLKLLAKELEVPVVAISQLNRGPEQRTDKKPMLADLRESGCLTASTRILRADTGAEVAFGELMRSGERPMVWSLDERLRMVARPMINVFPSGRKEVFRLRLASGREVEATGSHPFMKFEGWTPLAQLKVGDRIAAPRRVPEPIDTQRMPESELISLARMIGDGSCLKNQPIRYEPVDEANLAAVTVSAAHSDRAAIRDDYLAARVPSLRPARQRLPRGRCTPIAAWLAGLGLFTKRSHEKCVPEAVFRAPNDQVALFLRHLWSAGGSVRWDPTNGQGRVYYGSTSRRLIDDVAQLLLRVGIFSWITHAPKLGGHDSWRLHIHGAKDQVRFLRHVGVHGAEAVAAQEMLRQLKGPVRNPNLDSAPKKVWAQVRNRLSAKQMMDIQLHEPTMWKHSPSRSRPHRAEARIEDRAIHELARGDAYWDTVVEITSIGDQHVFDGTVSGTHNFVANGISLHNSLEQDADVVILLHRPDAFDRDDPRGGEADFILAKHRNGPTKTVTVAHQLHLSRFANMAR 894155A86DCB9D70 ssb_myctu Single-stranded DNA-binding protein ssb MTCY21D4.17 Rv0054 protein protein myctu Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv) MAGDTTITIVGNLTADPELRFTPSGAAVANFTVASTPRIYDRQTGEWKDGEALFLRCNIWREAAENVAESLTRGARVIVSGRLKQRSFETREGEKRTVIEVEVDEIGPSLRYATAKVNKASRSGGFGSGSRPAPAQTSSASGDDPWGSAPASGSFGGGDDEPPF 14797F8379FC2036 dnab-ssb 1 MtbSSB 5 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role bait bait over-expressed over expressed level over-expressed living cell living cell binding region binding region binding-associated region binding region deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain MtbDnaB 4 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role prey prey over-expressed over expressed level over-expressed living cell living cell tag fusion protein fusion protein undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association f4a Neither F21L nor N12S/P132L mutations affected the interaction of MtbDnaB and MtbSSB as the co-transformants containing F21L/MtbDnaB. (N12S/P132L)/MtbDnaB grew well on the screening medium. Furthermore, 10 amino acid residues from the SSB C-terminal, which affect the interaction between PriA and SSB in E. coli, were deleted from MtbSSB. Co-transformants containing SSBΔ10/MtbDnaB also grew well on the screening medium, but no growth was found when 20 amino acid residues were deleted from the MtbSSB C-terminal. This result suggests that the key interaction sites in MtbSSB are within the next 10 amino acid residues. These results also demonstrate specific interaction between MtbDnaB and MtbSSB. ssb-dnab 2 MtbSSB 5 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role prey prey purified purified binding region binding region binding-associated region binding region deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain gst tag gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined MtbDnaB 4 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role bait bait purified purified his tag his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined direct interaction direct interaction f4b The interaction of MtbDnaB with MtbSSB was further confirmed by surface plasmon resonance (SPR) assay. His-tagged MtbDnaB proteins were first immobilized on a nitrilotriacetate chip. As shown in Fig. 4B, significant responses of approximately 140 and 250 response units (RU) were found when increasing amounts of 30 and 60 nM GST-tagged MtbDnaB protein were passed over the chip, respectively (Fig. 4B, left panel). Similar results were also observed for the interaction of F21L, N12S/P132L, or SSBΔC10 with MtbDnaB, but a smaller response (48 RU) was observed when SSBΔC20 was passed over the chip (Fig. 4B, right panel). No response was observed for GST alone or GST-1505c. ssb-dnab-1 3 MtbDnaB 4 western blot western blot Immuno blot Immuno blot western blot western blot Immuno blot Immuno blot unspecified role unspecified role bait bait over-expressed over expressed level over-expressed living cell living cell his tag his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined MtbSSB 5 western blot western blot Immuno blot Immuno blot western blot western blot Immuno blot Immuno blot unspecified role unspecified role prey prey over-expressed over expressed level over-expressed living cell living cell his tag his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association f4c