cDNA cloning and expression of Carica papaya prochymopapain isoforms in Escherichia coli

Isolation of cDNA from Jacaratia mexicana encoding a mexicain-like cysteine protease gene

“…The homology comparison showed that JmCP4 and JmCP5 have 83% and 85% sequence identity with mexicain (P84346), respectively. JmCP4 and JmCP5 sequences have an identity of 88% with each other, supporting the hypothesis of their identity as CP isoforms within the latex of J. mexicana, as has been reported for other proteases like chymopapain isoforms I to V (Taylor et al, 1999). Table 2 shows a summary of the percentages of identity and accession numbers.…”

“…CP gene identification has occurred in several plant sources, such as Mesembryanthemum crystallinum (Forsthoefel et al, 1998), Daucus carota (Mitsuhashi et al, 2004), and C. candamarcensis (Pereira et al, 2001). Over-expression of CP genes from plant sources, like C. papaya (Taylor et al, 1999), Ipomoea batatas (Chen et al, 2009;2010), Nicotiana tabacum (Beyene et al, 2006;Zhang et al, 2009), and C. candamarcensis (Corrêa et al, 2011), has also been reported and shown to be encouraging examples of the aims presented in this study. The main objective of this research was to identify the gene sequences coding for the proteases that comprise the latex of J. mexicana.…”

Native and Biotechnologically Engineered Plant Proteases with Industrial Applications

“…There are five similar but distinct clones coding for precursor chymopapains (Taylor et al 1999). All of these isoforms have a free cysteine residue at position 117 (mature chymopapain numbering) not involved in disulfide bond formation or in the active site, and this property does differentiate chymopapain from the rest of the latex proteases (Azarkan et al 2003).…”

Revisiting the enzymes stored in the laticifers of Carica papaya in the context of their possible participation in the plant defence mechanism

“…The other differs at only one position (T 214 I) and, distinctively, contains a 19-amino-acid-long Cterminal extension. Furthermore, at least five similar but distinct cDNAs code for chymopapain [54], among [73] as well as those conserved in the radiocrystallographic structures of the mature enzymes [56 -59] are also shown: residues that participate in i ladders and those that contribute to h helices are indicated by the italised letters E and H, respectively. References to original sequencing work are given in the text.…”

“…E. coli invariably produces recombinant proproteinases entrapped in protein bodies from which they can be dissolved in the presence of chaotropic agents and subsequently refolded. In vitro conversion of the folded precursors into fully active proteinases can then proceed autocatalytically at pH 4 [52,54,66]. Activation of propapain appears to require several cleavage steps.…”