Composite organization of the cobalamin binding and cubilin recognition sites of intrinsic factor

Sergey N Fedosov, Natalya U Fedosova, Lars Berglund, Søren K Moestrup, Ebba Nexø, Torben E Petersen

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

Resumé

Intrinsic factor (IF(50)) is a cobalamin (Cbl)-transporting protein of 50 kDa, which can be cleaved into two fragments: the 30 kDa N-terminal peptide IF(30) and the 20 kDa C-terminal glycopeptide IF(20). Experiments on binding of Cbl to IF(30), IF(20), and IF(50) revealed comparable association rate constants (k(+)(Cbl) = 4 x 10(6), 14 x 10(6), and 26 x 10(6) M(-1) s(-1), respectively), but the equilibrium dissociation constants were essentially different (K(Cbl) = 200 microM, 0.2 microM, and <or=1 pM, respectively). The smaller fragment, IF(20), had unexpectedly high affinity for Cbl; however, efficient retention of the ligand required the presence of both fragments. Detailed schemes of the interaction of Cbl with IF(50) and with IF(30) and IF(20) are presented, where the sequential attachment of Cbl to the IF(20) and IF(30) domains plays the key role in recognition and retention of the ligand. Each isolated fragment of IF was tested for the binding to the specific receptor cubilin in the presence or absence of Cbl. Neither apo nor holo forms of IF(20) and IF(30) were recognized by the receptor. When two fragments were mixed and incubated with Cbl, they associated into a stable complex, IF(30+20).Cbl, which bound to cubilin as well as the noncleaved IF(50).Cbl complex. We suggest that formation of the cubilin recognition site on IF is caused by assembly of two distant domains, which allows the saturated protein to be recognized by the receptor. The obtained parameters for ligand and receptor binding indicate that both full-length IF(50) and the fragments may be involved in Cbl assimilation.

OriginalsprogEngelsk
TidsskriftBiochemistry
Vol/bind44
Udgave nummer9
Sider (fra-til)3604-14
Antal sider11
ISSN0006-2960
DOI
StatusUdgivet - 8. mar. 2005

Fingeraftryk

Intrinsic Factor
Vitamin B 12
Composite materials
Ligands
intrinsic factor-cobalamin receptor
Glycopeptides
Rate constants
Proteins

Citer dette

Fedosov, Sergey N ; Fedosova, Natalya U ; Berglund, Lars ; Moestrup, Søren K ; Nexø, Ebba ; Petersen, Torben E. / Composite organization of the cobalamin binding and cubilin recognition sites of intrinsic factor. I: Biochemistry. 2005 ; Bind 44, Nr. 9. s. 3604-14.
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Fedosov, SN, Fedosova, NU, Berglund, L, Moestrup, SK, Nexø, E & Petersen, TE 2005, 'Composite organization of the cobalamin binding and cubilin recognition sites of intrinsic factor', Biochemistry, bind 44, nr. 9, s. 3604-14. https://doi.org/10.1021/bi047936v

Composite organization of the cobalamin binding and cubilin recognition sites of intrinsic factor. / Fedosov, Sergey N; Fedosova, Natalya U; Berglund, Lars; Moestrup, Søren K; Nexø, Ebba; Petersen, Torben E.

I: Biochemistry, Bind 44, Nr. 9, 08.03.2005, s. 3604-14.

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

TY - JOUR

T1 - Composite organization of the cobalamin binding and cubilin recognition sites of intrinsic factor

AU - Fedosov, Sergey N

AU - Fedosova, Natalya U

AU - Berglund, Lars

AU - Moestrup, Søren K

AU - Nexø, Ebba

AU - Petersen, Torben E

PY - 2005/3/8

Y1 - 2005/3/8

N2 - Intrinsic factor (IF(50)) is a cobalamin (Cbl)-transporting protein of 50 kDa, which can be cleaved into two fragments: the 30 kDa N-terminal peptide IF(30) and the 20 kDa C-terminal glycopeptide IF(20). Experiments on binding of Cbl to IF(30), IF(20), and IF(50) revealed comparable association rate constants (k(+)(Cbl) = 4 x 10(6), 14 x 10(6), and 26 x 10(6) M(-1) s(-1), respectively), but the equilibrium dissociation constants were essentially different (K(Cbl) = 200 microM, 0.2 microM, and

AB - Intrinsic factor (IF(50)) is a cobalamin (Cbl)-transporting protein of 50 kDa, which can be cleaved into two fragments: the 30 kDa N-terminal peptide IF(30) and the 20 kDa C-terminal glycopeptide IF(20). Experiments on binding of Cbl to IF(30), IF(20), and IF(50) revealed comparable association rate constants (k(+)(Cbl) = 4 x 10(6), 14 x 10(6), and 26 x 10(6) M(-1) s(-1), respectively), but the equilibrium dissociation constants were essentially different (K(Cbl) = 200 microM, 0.2 microM, and

KW - Binding Sites

KW - Cobalt Radioisotopes

KW - Humans

KW - Intrinsic Factor

KW - Kinetics

KW - Peptide Fragments

KW - Protein Binding

KW - Protein Processing, Post-Translational

KW - Receptors, Cell Surface

KW - Recombinant Proteins

KW - Spectrometry, Fluorescence

KW - Surface Plasmon Resonance

KW - Vitamin B 12

U2 - 10.1021/bi047936v

DO - 10.1021/bi047936v

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SP - 3604

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JO - Biochemistry

JF - Biochemistry

SN - 0006-2960

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