Characterization of the mouse carboxypeptidase N small active subunit gene structure

Matthews, K.W.; Kildsgaard, Jens; Haviland, David L.; Wetsel, Rick A.

doi:10.1016/s0162-3109(00)80035-1

Cited by 2 publications

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“…33,35 It comprises two 83-kDa regulatory subunits (CPN2) and two approximately 50-kDa catalytic subunits (CPN1), which form a stable tetramer. [35][36][37][38] CPN was originally identified in plasma as the enzyme responsible for inactivating bradykinin and kallidin II by removal of their C-terminal arginine. 39 When cleaved by CPN, bradykinin and kallidin II change their receptor specificity, from B2 to B1, which effectively reduces their activity.…”

Section: Discussionmentioning

confidence: 99%

Identification of carboxypeptidase N as an enzyme responsible for C-terminal cleavage of stromal cell-derived factor-1α in the circulation

Davis

Singer

Sierra

et al. 2005

Blood

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The chemokine stromal-derived factor-1␣ (SDF-1␣) is an essential regulator of hematopoiesis, lymphocyte homing, pre-Bcell growth, and angiogenesis. As SDF-1␣ is constitutively expressed in many tissues, chemokine function is mostly regulated by proteolytic degradation. Human serum cleaves the 68-amino acid chemokine, SDF-1␣, at both termini. The enzyme or enzymes responsible for the removal of the carboxy-terminal lysine from SDF-1␣, leading to significant reduction in biologic activity, have not been identified.Using a new biochemical assay for measuring the carboxy-terminal cleavage activity, we purified from serum and plasma a peptidase that specifically removes the carboxy-terminal lysine from SDF-1␣ and identified it as carboxypeptidase N (CPN, also known as kininase I, arginine carboxypeptidase, and anaphylotoxin inactivator). We demonstrate that SDF-1␣ in serum and plasma lacks the carboxy terminal lysine, and depletion of CPN from serum and plasma significantly reduces the SDF-1␣ carboxypeptidase activity. Purified CPN effectively and specifically removes the carboxy-terminal lysine from SDF-1␣ and significantly reduces the chemokine's biologic activity as a pre-B-cell growth factor and chemoattractant. Thus, in addition to its role as a regulator of the biologic activity of kinins and anaphylatoxins, CPN is an important regulator of the biologic activity of SDF-1␣ by reducing the chemokine-specific activity. IntroductionThe chemokine stromal-derived factor-1 (SDF-1) and its unique receptor, CXCR4, are key regulators of the hematopoietic, nervous, and cardiovascular systems during embryogenesis. 1-3 Knock-out mice for SDF-1 or CXCR4 die perinatally with a similar phenotype, including defects in the formation of the cerebellum, the ventricular septum of the heart, the vasculature of the gastrointestinal system, and the lympho-hematopoietic system. [4][5][6][7] After birth, SDF-1 and CXCR4 continue to play key roles in regulating physiologic hematopoiesis and angiogenesis. [8][9][10][11][12][13][14][15] Studies in vitro have shown that SDF-1 functions as a growth factor for immature B lymphocytes and promotes chemotaxis in CXCR4-expressing cells, including tumor cells. 1,3,9,[16][17][18] Since CXCR4 can function as a coreceptor for T-cell tropic HIV-1, SDF-1 and CXCR4 may also affect the course of HIV-1 infection. 19,20 SDF-1␣ is a highly conserved chemokine in evolution, only a single conservative substitution (I to V) distinguishes the human chemokine from the mouse chemokine. 21,22 Two forms of SDF-1 are generated, ␣ and ␤, which are identical in amino acid sequence except for the presence of 4 additional amino acids at the carboxy terminus of SDF-1␤. 21,22 A third form of SDF-1 was identified in rat, SDF-1␥, which is predicted to code for a protein identical to SDF-1␤, except for the insertion of 30 additional amino acids at the carboxy terminus. 23 Structure-function studies have identified the amino terminal region of SDF-1 as critical to CXCR4 receptor activation. 22,24 SDF-1 lacking the amino term...

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Section: Discussionmentioning

confidence: 99%

Identification of carboxypeptidase N as an enzyme responsible for C-terminal cleavage of stromal cell-derived factor-1α in the circulation

Davis

Singer

Sierra

et al. 2005

Blood

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“…Molecular cloning [15] revealed that the 50 kDa subunit has sequence similarities to other metallocarboxypeptidases [17,21,31,32] although the homology is much higher with the members of the regulatory subfamily (~38 -49%) than with those in the pancreatic CPA/B subfamily (~14 -19%). However, most of the active site residues are conserved among the metallocarboxypeptidases and all require zinc as cofactor for activity [17,21,31].…”

Section: Comparison To Related Carboxypeptidasesmentioning

confidence: 99%

Structure and function of human plasma carboxypeptidase N, the anaphylatoxin inactivator

Skidgel

Erdös

2007

International Immunopharmacology

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Human carboxypeptidase N (CPN) was discovered in the early 1960s as a plasma enzyme that inactivates bradykinin and was identified 8 years later as the major "anaphylatoxin inactivator" of blood. CPN plays in important role in protecting the body from excessive buildup of potentially deleterious peptides that normally act as local autocrine or paracrine hormones. This review summarizes the structure, enzymatic properties and function of this important human enzyme, including insights gained by the recent elucidation of the crystal structure of the CPN catalytic subunit and structural modeling of the non-catalytic regulatory 83 kDa subunit. We also discuss its physiological role in cleaving substrates such as kinins, anaphylatoxins, creatine kinase, plasminogen receptors, hemoglobin and stromal cell-derived factor-1α (SDF-1α). HistoryCarboxypeptidases were initially isolated from pancreatic extracts and so were associated with protein and peptide degradation in the digestive tract [1][2][3][4]. However, work beginning in the early 1960's by Erdös and Sloane [5] revealed the presence of a novel carboxypeptidase in the blood that plays a regulatory role by inactivating bradykinin via removal of its C-terminal Arg residue. (Subsequent research some 2 decades later led to the discovery of a second kinin B 1 receptor activated by the carboxypeptidase metabolite that is its endogenous agonist -see below). Initially it was assumed that the enzyme represents a circulating form of pancreatic carboxypeptidase B, but this was disproved by the characterization of its enzymatic properties [5][6][7][8], which was later borne out by biochemical studies, sequence analysis [9][10][11][12][13][14][15] and ultimately X-ray crystallography [16]. To differentiate it from the pancreatic enzyme, it was named carboxypeptidase N (CPN; EC 3.4.17.3) and was the first member of a family of carboxypeptidases now known as the regulatory or CPN/E subfamily of metallocarboxypeptidases [17][18][19]. It has since been re-discovered many times as other substrates were found and so has acquired aliases such as creatine kinase conversion factor, plasma carboxypeptidase B, arginine carboxypeptidase, lysine carboxypeptidase and protaminase [8,17,20,21]. The most notable was the finding by Bokisch and Müller-Eberhard in 1969 -70 [22,23] that the human plasma "anaphylatoxin inactivator" is identical with CPN. This finding explains our interest in the work of Dr. Tony Hugli, who has made many seminal contributions to the understanding of the structure and function of anaphylatoxins [24][25][26][27]. In Send Correspondence and Proofs To: Randal A. Skidgel, PhD, Dept. of Pharmacology (M/C 868), Univ. of Illinois College of Medicine, 835 S. Wolcott, Chicago, IL 60612, Phone: 312-996-9179, FAX: 312-996-1648, EMAIL: E-mail: rskidgel@uic.edu. Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript w...

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Characterization of the mouse carboxypeptidase N small active subunit gene structure

Cited by 2 publications

References 0 publications

Identification of carboxypeptidase N as an enzyme responsible for C-terminal cleavage of stromal cell-derived factor-1α in the circulation

Identification of carboxypeptidase N as an enzyme responsible for C-terminal cleavage of stromal cell-derived factor-1α in the circulation

Structure and function of human plasma carboxypeptidase N, the anaphylatoxin inactivator

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