Molecular characterization of centerin, a germinal centre cell serpin.
Centerin [SERPINA9/GCET1 (germinal centre B-cell-expressed transcript 1)] is a serpin (serine protease inhibitor) whose expression is restricted to germinal centre B-cells and lymphoid malignancies with germinal centre B-cell maturation. Expression of centerin, together with bcl-6 and GCET2, constitutes a germinal centre B-cell signature, which is associated with a good prognosis in diffuse large B-cell lymphomas, but the molecular basis for this remains to be elucidated. We report here the cloning, expression and molecular characterization of bacterial recombinant centerin. Biophysical studies demonstrated that centerin was able to undergo the 'stressed to relaxed' conformational change which is an absolute requirement for protease inhibitory activity. Kinetic analysis showed that centerin rapidly inhibited the serine protease trypsin (k(a)=1.9x10(5) M(-1) x s(-1)) and also demonstrated measurable inhibition of thrombin (k(a)=1.17x10(3) M(-1) x s(-1)) and plasmin (k(a)=1.92x10(3) M(-1) x s(-1)). Centerin also bound DNA and unfractionated heparin, although there was no functionally significant impact on the rate of inhibition. These results suggest that centerin is likely to function in vivo in the germinal centre as an efficient inhibitor of a trypsin-like protease.
Paterson
Melinda A
MA
Australian Centre for Blood Diseases, Monash University, Level 6 Burnet Tower, 89 Commercial Road, Melbourne 3004, VIC, Australia.
Horvath
Anita J
AJ
Pike
Robert N
RN
Coughlin
Paul B
PB
D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D005786:Gene Expression Regulation; D006639:Histidine; D006801:Humans; D007700:Kinetics; D009363:Neoplasm Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015843:Serpins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D009363:Neoplasm Proteins; C476825:SERPINA9 protein, human; D015843:Serpins; D006639:Histidine