PubMed Journals: Biochemistry

  Source:		PMID: 10828960

    		Biochemistry. 2000 May 30;39(21):6459-65.
			Monoclonal antibody light chain with prothrombinase

			Thiagarajan P(1), Dannenbring R, Matsuura
			K, Tramontano A, Gololobov G, Paul S.

			Author Information
			(1) Departments of Internal Medicine and
			Pathology and Laboratory Medicine, Center
			for Chemical Immunology, University of Texas-Houston
			Medical School, 77030, USA.

			Prothrombin is the precursor of thrombin,
			a central enzyme in coagulation. Autoantibodies
			to prothrombin are associated with thromboembolism,
			but the mechanisms by which the antibodies
			modulate the coagulation processes are not
			understood. We screened a panel of 34 monoclonal
			antibody light chains isolated from patients
			with multiple myeloma for prothrombinase
			activity by an electrophoresis method. Two
			light chains with the activity were identified,
			and one of the light chains was characterized
			further. The prothrombinase activity eluted
			from a gel-filtration column run in denaturing
			solvent (6 M guanidine hydrochloride) at
			the characteristic positions of the light
			chain dimer and monomer. A constant level
			of catalytic activity was observed across
			the width of the light chain monomer peak,
			assessed as the cleavage of
			IEGR-methylcoumarinamide, a peptide substrate
			corresponding to residues 268-271 of prothrombin.
			Hydrolysis of this peptide by the light
			chain was saturable and consistent with
			Michaelis-Menten-Henri kinetics (K(m) 103
			microM; k(cat) of 2.62 x 10(-)(2)/min).
			Four cleavage sites in prothrombin were
			identified by N-terminal sequencing of the fragments:
			Arg(155)-Ser(156), Arg(271)-Thr(272),
			Arg(284)-Thr(285), and Arg(393)-Ser(394).
			The light chain did not cleave radiolabeled
			albumin, thyroglobulin, and annexin V under
			conditions that readily permitted detectable
			prothrombin cleavage. Two prothrombin fragments
			(M(r) 55 000 and 38 000), were isolated
			by anion-exchange chromatography and were
			observed to cleave a thrombin substrate,
			tosyl-GPR-nitroanilide. Conversion of fibrinogen
			to fibrin was accelerated by the prothrombin
			fragments generated by the light chain.
			These finding suggest a novel mechanism
			whereby antibodies can induce a procoagulant
			state, i.e., prothrombin activation via
			cleavage of the molecule.

			PMID: 10828960 [Indexed for MEDLINE]

     			                         Tweet       Print