Pectin, an important structural element in plants, is a group of acidic heteropolysaccharides composed of main chains of partially methyl-esterified α-1,4-linked D-galacturonic acid residues occasionally substituted at the C-2 or C-3 positions with one molecule of sugars such as L-rhamnose, D-galactose or D-xylose [1]. Pectinases are enzymes that decompose pectic substances, and are widely distributed in microorganisms and higher plants. According to their modes of action, pectinases can be divided into three categories: protopectinase, pectin esterase and pectin depolymerases (hydrolyase and lyase) [2]. The pectin depolymerase pectate lyase (Pel, EC 4.2.2.2) randomly cleaves the α-1,4-galacturonosidic bond of polygalacturonic acid (PGA) via a trans-elimination mechanism, and generates an unsaturated bond between C4 and C5 of the newly formed oligogalacturonide at the non-reducing end [3]. On the basis of amino acid sequences, Pels are classified into polysaccharide lyase (PL) families 1, 2, 3, 9 and 10 [4].

Ramie fibers are among the strongest and longest plant fibers, and are widely used in clothing fabrics, industrial packaging and cordage. However, the high content of non-cellulosic matter (20–35%, mainly pectin and hemicellulose) in raw ramie bast fiber hinders degumming [5]. To remove gum, highly alkaline treatment is performed in the traditional degumming process; this treatment partially damages the fibers, causes environmental problems by generating alkali-treated waste water and increases production costs by using high alkali doses. Pel degumming before NaOH treatment not only greatly improves the degumming effect but also decreases the amount of NaOH required, thus diminishing the drawbacks associated with NaOH use [6]. For the combined process with Pel degumming demand, Pel active in mildly alkaline environments with high specific enzymatic activity is preferred [7]. In previous studies, alkaline Pels derived from different microorganisms have been reported, such as from Bacillus Genus [3], [7], [8], Paenibacillus polymyxa [9], Dickeya dadantii [10], Klebsiella sp. [11] and Aspergillus nidulans [1], [12]. However, the low specific activity of these Pels requires use of large amounts of enzymes, thus increasing costs.

Ca2+ has a favorable effect on promoting the enzymatic activity of Pel [7]. Thus, addition of Ca2+ in the degumming process decreases the use of Pel. Various Pels have been described, for which the promoting effects of Ca2+ are insufficient (less than 10-fold increase in enzymatic activity). For example, the enzymatic activity of PEL1 from Massilia eurypsychrophila [13], Pel S6 from B. amyloliquefaciens S6 [14], BspPel from Bacillus sp. RN1 [15] and PelB from the soil metagenome [16] increases by 124%, 473%, 496.1% and 550%, respectively, with 0.5 mM Ca2+; the enzymatic activity of Pel from B. tequilensis SV11 [17], Pel DKS1 from B. pumilus DKS1 [18] and Pel4J4 from Dickeya dadantii DCE-01 [10] increases by 115.5%, 168% and 230.7%, respectively, with 1.0 mM Ca2+; and the enzymatic activity of PEL168P from B. subtilis [19] and PpPel9a from P. polymyxa KF-1 [4] increases by 138.3% and 166.5%, respectively, with 5.0 mM Ca2+. Additionally, reports of Ca2+ enhancement of the thermostability of Pel are scarce. Thus, a Pel with high specific enzymatic activity in mildly alkaline environments, whose enzymatic activity and thermostability are strongly promoted by Ca2+ is desirable in the textile industry.

In this study, a gene encoding a putative Pel was cloned from Bacillus sp. B58–2 and heterologously expressed in Escherichia coli. After purification through an Ni-NTA column, the recombinant Pel rBvelPL1-Ec was biochemically characterized and used for ramie fiber degumming. rBvelPL1-Ec had a high specific activity, and its enzymatic activity and thermostability were strongly promoted by Ca2+; thus, this enzyme may be valuable in the ramie degumming industry.