Proprotein Convertases Pcs

To date, seven mammalian members of subtilisin-related PCs that process substrates at basic residues have been identified. These include Furin/PACE, PC1/PC3, PC2, PC4, PACE4, PC5/PC6, and PC7/LPC/PC8/SPC7 (Figure 1).

This somewhat confusing nomenclature arose from the simultaneous discovery of some of these enzymes by different groups. PCs are multi-domain serine proteinases

A-Majid Khatib (ed.), Regulation of Carcinogenesis, Angiogenesis and Metastasis by the Proprotein Convertases, 7-26. © 2006 Springer.

Furin

PACE4

D H N S
D H N S

II II

Subtilisin

Kexin

Signal peptide Cysteine-rich domain Cytoplasmic domain

—. Pro-segment ■ Ser/Thr-rich domain RGD (S) sequence

Catalytic domain Transmembrane domain P domain

Figure 1. Schematic representation of the prohormone convertases PC1, PC2, Furin, PACE4, PC4, PC5 (A and B isoforms) and PC7. These PCs are multi-domain serine proteinases consisting of a signal peptide followed by prosegment, catalytic, middle, and cytoplasmic domains. Homology is highest in the catalytic domains and lowest in the carboxyl-terminal domains. The schematic representation for Kexin and subtilisin are given for comparison

Convertases Amino acid number Autocatalytic site Accession number

Furin

794

101A-K-R-R-T-K-R-D

NP_002560

PC1

751

105k-e-r-s-k-r-s-v

P21662

PC2

638

103g-f-d-r-k-k-r-g

P16519

PACE4

969

141q-e-v-k-r-r-v-k

P29122

PC4

654

105r-r-r-v-k-r-s-l

A54306

PC5

1870

109v-k-k-r-t-k-r-d

Q04592

PC7

785

134r-l-l-r-r-a-k-r

NP_004707

SKI-1

1052

131k-v-f-r-s-l-k-y

NP_003782

NARC-1

692

145e-d-s-s-v-f-a-q

NM_174936

Figure 2. Amino acid sequences of the autocatalytic sites of the PCs. Like their substrates, the prosegments of the PCs are removed at sites cleaved by the PCs. Indicated are the number of amino acid and accession number for every PC

Figure 2. Amino acid sequences of the autocatalytic sites of the PCs. Like their substrates, the prosegments of the PCs are removed at sites cleaved by the PCs. Indicated are the number of amino acid and accession number for every PC

consisting of a signal peptide followed by prosegment, catalytic, middle, and cytoplasmic domains (Figure 1). Homology is highest in the catalytic domains and lowest in the carboxyl-terminal domains.

These enzymes cleave precursor proteins at basic residues within the general motif (K/R)-(X)n-(K/R) j, where n = 0, 2,4 or 6 and X any amino acid except Cys [1-4]. Usually most of the PCs cleave their substrates at pairs of basic amino acids, but several of them, with monobasic sites are also cleaved [1-4]. Some PCs, such as PC1, PC2 and PC5A, are sorted and activated in the regulated secretory pathway and thus process protein precursors whose secretion is regulated. In contrast, the transmembrane proteins Furin, PACE4, PC5B and PC7 (Figure 1), cycle between the cell surface and the trans Golgi Network (TGN) and are involved in the processing of precursor proteins in the constitutive secretory pathway [1-4]. Like their substrates, the pro-segments of the PCs are also removed at a cleavage site containing a basic-amino acid PC motif (Figure 2), befitting their autoactivation [1-4].

1.1 Furin

Furin was the first convertase identified. Its discovery was made just after the availability of the Kex2 cDNA sequence. Kex2 is a cellular processing endopro-tease that is required for cleavage at dibasic sites within the killer toxin and the mating pheromone, a-factor precursors [5, 6]. In 1989, in an effort to find other

10

CHAPTER 1

Convertases/ Inhibitors

Human

Mouse

Chromosomes Cytogenetic

Chromosomes

Cytogenetic

Furin

15

15q26.1

7

7 D1-E2

PC1

5

5q15-q21

13

13C2

PC2

20

20p11.2

2

2G1

PACE4

15

15q26

7

7B5

PC4

19

19p13.3

10

10C1

PC5

9

9q21.3

19

19B

PC7

11

11q23.3

9

9A5.2

SKI-1

16

16q24

8

8E1

NARC-1

1

1p32.3

4

4C7

7B2

15

15q13-q14

2

2E5

Prosaas

X

Xp11.23

X

XA1.1

Figure 3. Chromosome localisation of mouse and human PCs and the inhibitors Prosaas and 7B2. Note the approximate position of PACE4 gene to fur gene on the human chromosome 15 and mouse chromosome 7 suggesting their probable common ancestry by gene duplication

Figure 3. Chromosome localisation of mouse and human PCs and the inhibitors Prosaas and 7B2. Note the approximate position of PACE4 gene to fur gene on the human chromosome 15 and mouse chromosome 7 suggesting their probable common ancestry by gene duplication related Kex2 enzymes, Fuller et al., identified the first mammalian homologue of Kex2 [7], fur gene. This gene is located on the human chromosome 15 and on mouse chromosome 7 (Figure 3). The Furin gene (PCSK3) was unexpectedly discovered by Roebroek et al., a few years earlier due to its proximity to the c-fes/fps protooncogene (fur being: c-fes/fps upstream region) [8]. At that time the product of the fur gene was believed to be a growth factor receptor because of the presence of a cysteine-rich domain and a putative trans-membrane domain in its sequence (Figure 1, [9]). Subsequently, the cloning of full-length Furin cDNA revealed that Furin was structurally analogous to Kex2, although the Ser/Thr-rich domain in Kex2 was replaced by a cysteine-rich domain (Figure 1, [10]). Furin is a membrane protein, initially produced as a 104 kDa pro-furin precursor which is rapidly converted into a 98 kDa form by an autocatalytic process (Figure 2, [ L1, 12].) This autocatalytic cleavage of the pro-Furin occurs in the endoplasmic reticulum (ER) and is a perquisite for the exit of the mature Furin molecule out of the ER to reach the cell surface [13, 14]. Unlike most other convertases, Furin has a widespread distribution being present in all tissues and cells examined so far.

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