Covalent binding of C3b to C4b within the classical complement pathway C5 convertase. Determination of amino acid residues involved in ester linkage formation.

Y.U Kim,M.C Carroll,D.E Isenman,M Nonaka,P Pramoonjago,J Takeda,K Inoue,T Kinoshita

doi:10.1016/s0021-9258(19)50644-5

Y.U Kim, M.C Carroll + Show 6 more

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https://doi.org/10.1016/s0021-9258(19)50644-5

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Journal: The Journal of biological chemistry	Publication Date: Feb 1, 1992
Citations: 61	License type: cc-by

Affiliation: Osaka University

Abstract

C5 convertase of the classical complement pathway is a protein complex consisting of C4b, C2a, and C3b. Within this complex C3b binds to C4b via an ester linkage. We now present evidence that the covalent C3b-binding site on human C4b is Ser at position 1217 of C4. We also show that formation of the covalently linked C4b.C3b complex occurs in the mouse complement system and that the C3b-binding site on mouse C4b is Ser at position 1213 which is homologous to Ser-1217 of human C4. Therefore, covalent binding of C3b to a single specific site on C4b within the classical pathway C5 convertase is likely a common phenomenon in the mammalian complement system. Specific noncovalent association of metastable C3b with C4b would occur first, leading to reaction of the thioester with a specific hydroxy group. This is supported by two lines of experimental evidence, one which shows that a mutant C4 that does not make a covalent linkage with C3b is still capable of forming C5 convertase and a second in which the C4b.C3b complex has been demonstrated by cross-linking erythrocytes bearing this C5 convertase.

Highlights

From the $Departmentof Bacteriology, Osaka University Medical School, Osaka 565, Japan, the §Departmentof Pathology, Haruard Medical School, Boston, Massachusetts 02138, the TlDepartment of Biochemistry, Uniuersity of Toronto, Toronto, M5S IAl Ontario, Canada, the IlDepartment of Immunobiology, Cancer Research Institute, Kanazawa Uniuersity, Kanazawa920, Japan, and the **Department of Immunoregulation, Research Institute for Microbial Diseases, Osaka Uniuersity, Osaka 565, Japan
We show that formation of the covalently linked C4beC3bcomplex occurs in the mouse complement system and that the C3b-binding site on mouse C4b is Ser at position 1213 which is homologous to Ser-1217 of human C4
One of the C3b molecules binds t o C3 convertase itself, forming a trimolecular complex C4b2a3b [8].This trimolecular complex is the C5 convertase in which both C3b and C4b participate in C5 binding [8,9,10,11,12]

Summary

DETERMINATION OF AMINO ACID RESIDUESINVOLVEDINESTERLINKAGEFORMATION*

C5 convertase of the classical complement pathway IgG or IgM, an enzyme that convertsC3 to C3b is formed on is a protein complex consisting of C4b, C2a, and C3b Within this complex C3b binds toC4b via an ester linkage. Specific noncovalent association of metastable C3b with C4b would occur first, leading to reaction of the thioester with a specific hydroxy group This is supportedby two lines of experimental evidence, one which shows that a mutant C4that does not make a covalent linkage the targetsurface [1,2]. We demonstrated the important characteristicof this component is the intramolec-formation of a noncovalently associatedtrimolecular C4b2a3b ularthioesterbondformed between a sulfhydryl group of complex with C5 convertase activitywhen the covalent bindcysteine and a carbonyl group of glutamic acid [3, 4]

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