Technical note Purification of α1-antichymotrypsin from human plasma with recombinant M. catarrhalis ubiquitous surface protein A1 ☆ Taras Manolov, Arne Forsgren, Kristian Riesbeck ⁎ Medical Microbiology, Department of Laboratory Medicine, Malmö University Hospital, Lund University, SE-205 02 Malmö, Sweden Received 21 March 2007; received in revised form 6 December 2007; accepted 12 December 2007 Available online 15 January 2008 Abstract Alpha 1-antichymotrypsin (ACT) inhibits chymotrypsin-like enzymes, particularly neutrophil cathepsin G. Moreover, ACT in its native form suppresses chemotaxis of neutrophils and decreases neutrophil production of superoxide radicals. We recently showed that Moraxella catarrhalis ubiquitous surface protein (Usp) A1 is able to specifically bind ACT in the context of a novel virulence mechanism. In this study, we report that recombinant UspA1 557–704 coupled to CNBr-Sepharose can be used in a simple one-step purification of ACT from human plasma. UspA1 557–704 -purified ACT remains intact and active as shown by binding to M. catarrhalis and a chymotrypsin inhibition assay. The novel method for ACT isolation from plasma has important advantages in simplicity and time as compared to conventional methods. © 2007 Elsevier B.V. All rights reserved. Keywords: Antichymotrypsin; Moraxella catarrhalis; Purification; UspA1 1. Introduction α1-Antichymotrypsin (ACT) is a member of the serine proteinase inhibitor gene family and is a glycoprotein with a molecular mass of 65 kDa, of which approximately 25% are oligosaccharides (Travis and Salvesen, 1983). ACT is an inhibitor of chymotrypsin-like serine proteases, and the primary target is considered to be neutrophil cathepsin G(Beatty et al., 1980). ACT is mainly produced by hepatocytes and excreted into the bloodstream. After tissue damage the plasma concentration of ACT may double within 8 h, and it is therefore regarded as an acute phase reactant. At a site of infection ACT functions as an inhibitor of neutrophil proteases, mainly cathepsin G. Interestingly, the rate of cathepsin G inhibition with ACT is at least 1000-fold higher than inhibition of any other chymotrypsin-like enzyme (Travis and Salvesen, 1983). When bound to its target enzymes, ACT is also a chemoattractant for neutrophils. In contrast, native ACT has a suppressive effect on neutrophil chemotaxis. Both native and target enzyme-bound ACT is capable of inhibiting superoxide production by neutrophils. Morover, ACT is known to inhibit tumour necrosis factor (TNF)-α-induced platelet-activating factor (PAF) synthesis in neutrophils, macrophages, and endothelial cells (Camussi et al., 1988). Journal of Immunological Methods 333 (2008) 180 – 185 www.elsevier.com/locate/jim ☆ This work was supported by grants from the Alfred Österlund, the Anna and Edwin Berger, the Crafoord, the Greta and Johan Kock Foundations, the Swedish Medical Research Council, the Swedish Society of Medicine, and the Cancer Foundation at the University Hospital in Malmö. ⁎ Corresponding author. Tel.: +46 40 338494; fax: +46 40 336234. E-mail address: kristian.riesbeck@med.lu.se (K. Riesbeck). 0022-1759/$ - see front matter © 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.jim.2007.12.012