Alginate Lyases from Marine Bacteria: An Enzyme Ocean for Sustainable Future

The cell wall of brown algae contains alginate as a major constituent. This anionic polymer is a composite of β-d-mannuronate (M) and α-l-guluronate (G). Alginate can be degraded into oligosaccharides; both the polymer and its products exhibit antioxidative, antimicrobial, and immunomodulatory activities and, hence, find many commercial applications. Alginate is attacked by various enzymes, collectively termed alginate lyases, that degrade glycosidic bonds through β-elimination. Considering the abundance of brown algae in marine ecosystems, alginate is an important source of nutrients for marine organisms, and therefore, alginate lyases play a significant role in marine carbon recycling. Various marine microorganisms, particularly those that thrive in association with brown algae, have been reported as producers of alginate lyases. Conceivably, the marine-derived alginate lyases demonstrate salt tolerance, and many are activated in the presence of salts and, therefore, find applications in the food industry. Therefore, this review summarizes the structural and biochemical features of marine bacterial alginate lyases along with their applications. This comprehensive information can aid in the expansion of future prospects of alginate lyases.

Primarily, brown algae have a complex sugar composition, mainly including mannitol, laminarin, and alginate [17]. Mannitol is an alcohol derived from mannose, whereas laminarin is a polymer of β-1,3-linked glucose residues branched at 1,6-b [18,19]. Mannitol and laminarin are carbohydrate reserves that are accumulated by the algae during summer, and

Alginate Lyase-Producing Marine Bacteria
Large quantities of alginates are produced by various algae in the ocean every year, they serve as nutrient resources for heterotrophic marine bacteria and, thus, play an ecological role in coastal ecosystems, similar to that of cellulosic and hemicellulosic biomass in terrestrial environments. Various alginate lyases produced by marine microbes play important roles in marine alginate degradation. A couple of alginate lyases were separated from different kinds of microorganisms in the past several years, especially from the bacteria on brown algae (such as Bacillus sp. obtained from rotten seaweed) [88], Paenibacillus algicola isolated from rotten brown algae samples collected from China [92], and Pseudoalteromonas sp. SM0524 separated from marine kelp residues [93]. Alginatedegrading bacteria were screened and identified from brown algae collected from a French beach and the Arctic region, which belonged to the classes Gamma-proteobacteria and Flavobacteria of the phylum Proteobacteria and Bacteroidetes [94,95]. Wang et al. (2017) reported that 12 different bacterial strains belonging to eight genera were recovered from the three brown algae (Laminaria japonica, Sargassum horneri and Sargassum siliquastrum) samples obtained from the coast of Nanhuangcheng Island, China, capable of excreting alginate lyases [25]. In addition, an alginate lyase-producing bacteria Vibrio. sp. QD-5 was isolated from rotten kelp [96]. Strain BP-2 producing the alginate lyase was screened and identified from rotted Sargassum collected from Weizhou Island, China [97].

Enzymatic Properties of Alginate Lyases from Marine Bacteria
Most of the marine-based alginate lyases are endolytic enzymes, which could break down glycosidic bonds of alginate and thus produce unsaturated oligosaccharides (Table 1). Endolytic alginate lyases were employed to prepare AOSs with various DPs. For example, Swift et al. discovered an endo-type alginate lyase AlgMsp from a marine bacterium Microbulbifer sp. 6532A, which produces AOSs DP2-5 [46]. Alg7D, an endo-type alginate lyase separated from Saccharophagus degradans 2-40 T mainly produced oligosaccharides with a DP of 3-5 [98]. It has been disclosed that depolymerized low DP alginate prepared through an enzymatic converter possesses various kinds of biological activities [63,99]. Nguyen et al. prepared a series of AOSs with the potential for efficient production of low DP alginate oligosaccharides by using a new marine actinobacterium-produced alginate lyase AlyDS44 Streptomyces luridiscabiei [100]. In addition, Aly-IV from Vibrio. sp. QD-5 [96] and AlgA from Pseudomonas sp. E03 [101] are two novel endolytic alginate lyase enzymes that can release a range of AOSs with low DP. In addition, a few exolytic alginate lyases could directly monomerize alginate to a monosaccharide [102] (Table 1). Interestingly, novel alginate lyases isolated from Microbulbifer sp. SH-1 [103] and BP-2 strain [97] demonstrated both exolytic and endolytic cleavage activities.
Substrate-specific alginate lyases are able to be utilized for determining sequences of alginate substrates and producing oligosaccharides with certain structures. The substrate specificities of these alginate lyases largely rely on their architectures, amino acid residues, and the alignment of the saccharide residues in the substrate. Various alginate lyases could recognize four different types of linkages, including G-G, M-M, G-M and M-G. The ALG-5 from Streptomyces sp. ALG-5 depolymerizes the polyG substrate [104]. The Alyw203 from Vibrio sp. W2 is also a polyG-specific alginate lyase [105]. High-alkaline alginate lyase, A1m, is a kind of mutant enzyme with cleavage specificity for the G-G linkage [91]. In addition, AlyPB2 from Photobacterium sp. FC615 specifically depolymerizes polyM [83]. However, there are several alginate lyases showing activities in both of them such as the lyases from Vibrio sp. QY108 [106], Cobetia sp. NAP1 [107], Pseudoalteromonas sp. SM0524 [93], Pseudoalteromonas carrageenovora ASY5 [108], Agarivorans sp. L11 [109], and Streptomyces luridiscabiei [100]. Moreover, bifunctional lyases possess different degradation activities toward different substrates. For instance, Aly-SJ02, a bifunctional alginate lyase from Pseudoalteromonas sp. SM0524, was preferable to depolymerizes poly (M) than poly (G) [93]. Aly-SJ02 showed lower K m to polyG than that of polyM and sodium alginate [93]. Belik et al. reported a bifunctional endolytic alginate lyasesALFA3isolated from Formosaalgae KMM 3553 T [110]. These studies suggested that the bifunctional alginate lyases in alginate-utilizing bacteria could provide an efficient mechanism to utilize rich and reliable alginate sources for producing energy. Glaciecolachathamensis S18K6T -polyG AlyGC -6 -- [116] Vibrio sp. W2 -polyG Alyw203 endo-type 7 1-2 - [105] According to the amino acid sequence and structural features, alginate lyases could be classified into several polysaccharide lyase (PL) families. As indicated in Table 1, marine bacteria-based alginate lyases are mainly PL6 and PL7 family members, which are endolytic. Moreover, alginate lyases are grouped into families based on the threedimensional structures, which makes it possible to research the relationship between structure and function. The parallel β-helix family includes VsAly7D from Vibrio sp. QY108 [106], which belongs to the PL-7 family and AlyGC from Glaciecola chathamensis S18K6T [116], which belongs to the PL6 family, while the jelly-roll family includes Aly-SJ02 from Pseudoalteromonas sp. SM0524 of PL18 [117] and AlyA5 and AlyA1 from Zobellia galactanivorans of the PL-7 family [90].
Notably, some alginate-degrading strains could produce several alginate lyases to synergistically degrade exogenous alginate. The Pseudoalteromonas sp. strain ASY5 generates two extracellular alginate lyases Alg823 and Aly1281 (Table 1), which have similar action mode and main degradation products but different specificities to substrate. Although Alg823 andAly1281 are both bifunctional, Alg823 demonstrates the highest activity with polyM [68], while Aly1281 shows higher activity with polyG than that of polyM [108]. The similar action modes and main degradation products may bring them maximum enzyme activity under the same environmental conditions, and the substrate specificity difference leads to a synergistic alginate degradation effect of Alg823 and Aly1281. Photobacterium sp. FC615 produces extracellular (AlyPB1) and intracellular (AlyPB2) alginate lyases. Two alginate lyases have different substrate specificities, families, and modes of action. AlyPB1 is an alginate lyase with a preference for polyG, and AlyPB2 is a bifunctional lyase [83]. Pseudoalteromonas sp. 0524 secrets two extracellular alginate lyases (AlyPM and Aly-SJ02), which have different substrate specificities and, thus, synergistically facilitate the alginate degradation [93,113]. Additionally, Formosa algae KMM 3553 T secretes two endolytic alginate lyases (ALFA3 and ALFA4) with different substrate specificities. ALFA3 is a bifunctional lyase, while ALFA4 degrades only mannuronate blocks [110]. Zobellia galactanivorans produce two intracellular alginate lyases (AlyA1PL 7 and AlyA5) with different modes of action [90].

Biochemical Properties of Marine Bacteria-Produced Alginate Lyases
There are some characteristics of alginate lyases produced from marine bacteria that are shown in Table 2. The optimal working conditions for most of the alginate lyases (especially the PL7 enzyme family) are between pH 7.0 and 8.5. Additionally, several alginate lyases exhibit the optimal activities in alkaline (Alyw203 from Vibrio sp. W2 [105]) and acidic (ALFA3 from Formosa algae KMM 3553 T [110] and SALy from Sphingomonas sp. [107]) environments (Table 2). Lyase Alyw202 has an optimal pH of 9.0, while the optimum pH value for lyases AlyM, AlyA1PL7, and AlyA5 is 7.0. The optimal pH of AlgMsp, AlyPB1, AlyPB2, ALG-5, AlgC-PL7, Aly1281, Alg823, and ALFA4 at pH 8.0 is between those values ( Table 2). In addition, VsAly7D from Vibrio sp. QY108 showed its maximum activity at a pH of 8.0, and the enzyme stability remained within the pH range of 8.0 to 10.0. Therefore, VsAly7D works as an alkaline-stable alginate lyase that is generally stored under weak alkaline conditions and adapts different environments [106]. AlyPM showed the maximum activity at pH 8.5 and maintained~70% of the maximum activity from pH 7.0 to 9.5 [113]. AlgC-PL7 retained~50% of its maximum activity from pH 6 to 9. These results indicated that AlgC-PL7 generally possesses optimal activity under neutral conditions [107]. AlySJ-02 from Pseudoalteromonas sp. SM0524 demonstrated maximal activity at pH 8.5 and retained >50% activity at pH 7.0-10 after 20 min incubation [93]. Cold-adapted alginate lyase AlyL1 from Agarivorans sp. L11 showed the highest activity at a pH of 8.6 and maintained its stability from a pH of 6.0 to 9.6 [109].

Enzyme Kinetics of Alginate Lyases from Marine Bacteria
Enzyme kinetics is an essential factor in evaluating the catalytic capability of an enzyme toward practical applications. However, since the alginate substrate is biochemically heterogeneous and alginates produced by various seaweeds have different mannuronic/guluronic (M/G) ratios, the enzyme kinetics of alginate lyases was difficult to measure. Additionally, the polyM, polyG, and polyMG subdomains and their frequencies are significantly different in different seaweed sources [60,122]. Alginate lyase-mediated production of alginate usually causes a mixture of polymers with different DP values, and their average length was determined by the preparation methodology and conditions. Therefore, it is hard to compare the enzyme kinetics among different alginate lyases [46]. The kinetic parameters of marine bacteria-based alginate lyases towards different substrates are shown in Table 3. For instance, with the substrate sodium alginate, the K m and V max of AlyH1 were measured as 2.28 mg/mL and 2.81 U/mg, respectively, indicating that AlyH1 (under sodium alginate substrate) possesses high enzyme efficiency [114]. Zhang et al. (2020) investigated the salt effect (NaCl: 300 mM; KCl: 1000 mM) on the enzyme kinetics of Aly1281 (substrate: sodium alginate), and it was found that adding 300 and 1000 mM of NaCl could decrease the K m value by 54.9% and 74.3%, respectively. Compared to the K m values under electrolyte-free conditions, the result indicated that the affinity of substrate and catalytic activity of alginate lyases could be greatly enhanced by adding salts or electrolytes, which is the salt-activation effect [108]. AlgMsp from Microbulbifer sp. 6532A showed a K m of 3.4 mM for alginate [46]. Additionally, the catalytic efficiency (k cat /K m ) of AlyL1 to alginate was calculated as 9952.8 ± 33.1 mg mL −1 s −1 . AlyL1 exhibits a K m value of 0.19 ± 0.04 mg/mL with a V max value of 907.8 ± 72.5 U/mg protein. The results suggested that AlyL1 possesses a high affinity to alginate and could efficiently degrade alginates into oligosaccharides [109]. Moreover, K m values of AlyA1 (PL7 family) from Zobellia galactanivorans with various seaweed alginate substrates range from 1.7 to 6.2 mM, with increased binding affinity to alginate with higher guluronate composition [90]. In addition, Aly-SJ02, an alginate lyase from Pseudoalteromonas sp. SM0524, has a higher K m of 6.1 mM towards the alginate [93]. For seaweed-intake marine organisms, the low binding affinity of alginate lyases is acceptable due to the high concentration of alginate contents in seaweed (e.g., 17-45% w/w of dried brown seaweed) [21]. There are some notable exceptions of alginate lyases with K m values in the micromolar range. Alginate lyases from different marine sources could have different polyM, polyG, or polyMG substrate specificities [60]. Typically, some alginate lyases prefer one substrate but still cleave the other substrates at a reduced rate. For example, Aly-SJ02, an alginate lyase from Pseudoalteromonas sp SM0524, degrades polyG and polyM with polyG-specific activity and 75% of that against polyM [93]. Additionally, the K m and k cat /K m of VsAly7D to alginate were calculated as 0.217 mM and 227 L mol −1 s −1 , respectively [106]. Bifunctional alginate lyases could degrade different types of alginates, making them potential biocatalysts for industrial application.

Preparation of AOs
Alginate oligosaccharides (AOs) possess various biological properties that provide benefits for improving human health. Their bioactivities, including antitumor [128], antidiabetic [129], antihypertensive [130], anti-inflammatory [131,132], antimicrobial [133], antioxidant [134], anticancer [99], immunomodulatory [135,136] and anti-radiation [43,137] properties, have been comprehensively summarized. Generally, traditional preparation methods for the production of AOs are usually under strong acidic and alkaline conditions [138], thus resulting in severe environmental damage. In contrast, enzyme-based AOs production methods are more "green" and environmentally sustainable. AOs prepared by enzymatic degradation methods showed special bioactivities due to their unsaturated oligosaccharide structures [139,140]. However, there is only one commercially available alginate lyase (CAS number: 9024-15-1, Sigma-Aldrich, St. Louis, MO, USA) with a high pH tolerance, high catalytic activity (>10,000 U/g) and magnificent heat stability, which is expensive and only sold in the form of reagents, and most of the marine bacterial-produced alginate lyases are just investigated at the laboratory level. NitAly obtained from Nitratiruptor sp. SB155-2 shows the highest alginate lyase activity at 70 • C [141], while alginate lyase Aly-IV (PL7 family) from Vibrio. sp. QD-5 [96] and Aly08 from Vibrio sp. SY01 [142] are alkaline-stable, with optimal working pH values of 8.9 and 8.35, respectively.
Apart from the above-mentioned pH and thermo-stable alginate lyases, several alginate lyases demonstrated great potential for producing alginate oligomers with various DPs. Since the bioactivities of AOs are largely dependent on their DP values and chemical structures [143,144], endolytic alginate lyase-produced oligosaccharides with various DPs and diverse structures have attracted significant attention. The investigations of new AOsproducing alginate lyases were mostly conducted at the laboratory scale, and it could be seen that the endolytic alginate lyase generally produced alginate oligomers with DPs ranging from 2 to 5 [144]. For instance, the alginate lyase isolated from Isoptericola halotolerans CGMCC 5336, purified by gel column chromatography and characterized by TLC and ESI-MS, could perform an elimination reaction on guluronic acid (active sites: G or G-Gresidues) and generate oligomers with DPs of 2-4 [145] (Table 4). Alg2A, an endolytic alginate lyase from Flavobacterium sp. S20, can produce oligosaccharides with high yields along with high DP values (e.g., DP5 (penta-), DP6 (hexa-) and DP7 (hepta-)saccharides) [146] (Table 4). Zhu et al. degraded alginate with alginate lyase from Flammeovirga sp. NJ-04 to prepare oligosaccharides with DP2-4 [58] (Table 4).
Notably, the combination of some endolytic and exolytic lyases could lead to a remarkable synergistic effect on the degradation of alginate. For AOs preparation, the simultaneous application of endolytic lyase AlyPB1 and exolytic lyase AlyPB2 could lead to significantly increased conversion from alginate to unsaturated monosaccharides, which could reach approximately seven-fold that of single AlyPB2 [83] (Table 4). Moreover, substrate-specific alginate lyases could be employed for the preparation of oligosaccharides with a specific molecular structure. Anne et al. constructed a diguluronic acid linkage-cleavable alginate lyase, which could be employed for the preparation of guluronic acid oligosaccharide [147]. Zhu et al. isolated a novel polyM-specific alginate lyase AlgNJ-07 from Serratia marcescens NJ-07, which showed good PolyM-degradation efficiency [81] and thus could act as a potential tool for the production of mannuronic acid oligosaccharide (Table 4). Table 4. Some applications of alginate lyase from marine bacteria.

Enzyme
Source Application References Field of Application

ALFA3
Formosa algae KMM 3553 T Preparation of alginate oligosaccharides [110] in agriculture, in feed production, to lower cholesterol levels in blood plasma

Aly1281
Pseudoalteromonascarrageenovora ASY5 Preparation of alginate oligosaccharides [108] in agriculture, feed production AlgNJ-07 Serratia marcescens NJ-07 Preparation of alginate oligosaccharides [81] antimicrobials AlgNJ-07 Serratia marcescens NJ-07 Preparation of alginate oligosaccharides [81] antimicrobials for the treatment of cystic fibrosis, in agriculture, in feed production, in medicine for the diagnosis of diseases, to lower cholesterol in blood plasma

FsAlgB
Flammeovirga sp. NJ-04 Preparation of alginate oligosaccharides [58] in medicine for the diagnosis Aly Pseudomonas sp. HZJ 216 Preparation of alginate oligosaccharides [148] antimicrobials, in medicine for the diagnosis of diseases

Aly5
Flammeovirga sp. Strain MY04 Preparation of alginate oligosaccharides [149] in medicine for the diagnosis

AlyPB1 and AlyPB2
Photobacterium sp. FC615 Preparation of unsaturated monosaccharide [83] antimicrobials for the treatment of cystic fibrosis

Alg7A
Vibrio sp. W13 Preparation of alginate oligosaccharides [144] inhibition of lipid oxidation in food emulsions

AlyL1
Agarivorans sp. L11 Produce TPC for bioenergy production [151] inhibition of lipid oxidation in industrial emulsions

AlyPB2
Photobacterium sp. FC615 Alginate Sequencing [83] in the production of alginates Aly SM0524 Pseudoalteromonas sp. SM0524 Preparation of bioethanol [152] antimicrobials for the treatment of cystic fibrosis, for lowering plasma cholesterol levels

Anti-Biofilm Activity
It is difficult for normal antibiotics to kill some pathogenic bacteria with complex biofilms on their surfaces. It was disclosed that alginate components in the biofilm of Pseudomonas aeruginosa could protect them from being recognized and cleared by the immune system and resisting antibiotic treatment [124,155]. Therefore, using a purified alginate lyase-antibiotic complex to synergistically treat Pseudomonas aeruginosa infections is a possible therapeutic method [125,156]. Recently, a purified alginate lyase (AlyP1400) from a marine Pseudoalteromonas sp. 1400 bacterium demonstrated the capability of disrupting the formation of biofilms of Pseudomonas aeruginosa by decomposing alginate within the extracellular polysaccharide matrix and thus enhancing the bactericidal activity of tobramycin, which may act as a promising strategy for combinational therapy [150] (Table 4).

Bioethanol Production
The alginate lyases are also employed as a potential tool for producing bioethanol. The exo-type alginate lyase depolymerizes the alginate oligomers into unsaturated monosaccharides and subsequently non-enzymatically converted to 4-deoxy-L-erythro-hexoseulose uronic acid (DEH), which was then reduced into 2-keto-3-deoxy-gluconate (KDG) by DEH reductase and was further connected to the Entner-Doudoroff (ED) pathway [157]. Normally, industrial microorganisms cannot directly utilize alginate as a starting resource to produce ethanol due to the lack of an alginate-mediated metabolic pathway. For a long time, it has been difficult to achieve efficient production of ethanol from brown algae. In 2012, Wargacki et al. [152] designed and prepared a bio-ethanol synthesis microbial platform using E. coli as a producer to secrete alginate lyase SM0524Aly from Pseudoalteromonas sp. SM0524 by an auto transporter (Table 4). Additionally, in Vibrio splendidus 12B01, an alginate lyase-encoding large gene cluster was introduced along with alginate catabolismauxiliary gene clusters for achieving appropriate metabolism pathways. Finally, a pyruvate decarboxylase (Pdc) and an alcohol dehydrogenase B (AdhB)-encoding gene cluster was integrated into the E. coli chromosome to produce bioethanol. Moreover, endogenous E. coli genes, which encode fermentative byproducts, were removed. Accordingly, the fermentative yield of alginate, mannitol, and glucan could reach 0.28 g ethanol/per g dry brown algae (>80% of the maximum theoretical yield) [152]. Yagi et al. (2016) utilized Alg17C, an exo-oligoalginate lyase (PL7 family) isolated from halophilic Gram-negative bacterium Cobetia sp. NAP1 (brown algae Padina arborescens Holmes, as the bacterium resource) to depolymerize alginate into a monomeric sugar acid. Furthermore, Yagidis concluded that Alg17C could serve as the key enzyme to produce alginate monomers in the process of utilizing alginate for the production of biofuels and chemicals [107] (Table 4). It has been reported that the alginate lyase from Shewanella sp. Kz7 could degrade polyG blocks of alginate and accordingly produce monosaccharides such as 6-tetrahydroxy tetrahydro-2Hpyran-2-carboxylic acid (TPC), a useful intermediate for biofuel production [153] (Table 4).

Disposal of Seaweed Waste
In recent years, the amount of seaweed waste has drastically increased worldwide. One of the main organic components in seaweed is alginate, the content of which is as high as 50% in seaweed species such as wakame (Undaria pinnatifida) [158]. The disposal and re-utilization of seaweed waste are essential issues for the protection of marine environments and recycling of sustainable biomass. However, the degradation of alginate by general microorganisms is not easy to realize, mainly due to the complicated structures and molecular alignments of alginate. Thus, the isolation of specific microorganisms for alginate degradation is highly demanded, which is essential for the effective disposal of seaweed wastes. Tang et al. (2009) utilized alginate lyase-producing bacteria strain A7 (Gracilibacillus sp.) to degrade alginate in the wakame composting process. In a laboratory-scale test, after 72 h of composting, the alginate content in the wakame remarkably diminished from an initial value of 36.0% to 14.3%, suggesting the effectiveness of A7 for alginate decomposition [154] (Table 4).

Elucidate the Structure of Alginate
To profoundly understand the influence of the polymer architecture on the physicochemical properties of alginate, alginate lyases have been utilized to analyze the fine polymer architecture, especially the alignment of α-L-guluronate (G) and β-D-mannuronate (M) units of alginate. It is also very necessary to investigate the fine architecture of alginate for the preparation of tailor-made alginate. Lu et al. combined 1 H NMR spectroscopy with exolytic alginate lyase AlyPB2 to establish a method for sequencing alginate oligosaccharides [83] (Table 4). Compared with the traditional sequencing method, this method provides a simple strategy for characterizing the structure of alginate oligosaccharides.
When studying the mechanism of action of alginate lyases, it was found that most of the studied alginate lyases function endolytically, i.e., they split the alginate molecules from the inside and do not produce significant amounts of oligomers at the beginning of the reaction [48]. If the reaction proceeds, the end products are typically dimers, trimers, tetramers, or pentamers [85]. However, several exoliases were described that remove single residues from the polymer end [48,160].
Gacesa [161] was the first to propose a reaction mechanism for alginate lyases. First, the negative charge on the carboxylate anion is shielded by the enzyme. This allows the proton to be abstracted from C-5. It is proposed to stabilize the intermediate enolate ion by resonance. Finally, electron transfer from the carboxyl group results in the formation of a double bond between C-4 and C-5 and cleavage of the O-glycosidic bond. It was found that cleavage is promoted by an amino acid residue acting as an acid [162]. The new non-reducing end will contain 4-deoxy-L-erythro-hex-4-enepyranosyluronate (∆). This double bond is absorbed at 235 nm and is used to quantify alginate lyase activity [48]. The negative charge of most alginate lyases is stabilized by glutamine, arginine, or asparagine. It is important for the catalytic mechanism that, for M-residues, the C-5-proton and the departing oxygen on C-4 lie syn relative to each other, while for G residues they lie anti relative to each other. For the studied alginate lyases, it was found that for M-specific lyases, the C-5 proton is abstracted by tyrosine, which also acts as an acid facilitating the cleavage of the O-glycosidic bond. For lyases acting on G-residues, the C-5 proton is abstracted by histidine, while tyrosine again acts as an acid [162]. Alginate lyases belonging to the PL6 family do not follow this pattern. They use Ca 2+ as a neutralizer, lysine as a proton abstracting residue, and arginine as an acid [162].  [159]. (a) Alginate lyase from P. Algicola; (b) alginate lyase from L. Japonica; (c) alginate lyase from U. Pinnatifida; (d) alginate lyase from P. arborescens Holmes (e) alginate lyase from I. halotolerans.

Conclusions Remarks
Thus, each year, various algae in the ocean produce large amounts of alginates, which serve as nutrient resources for heterotrophic marine bacteria and thus play an ecological role in coastal ecosystems similar to that of cellulose and hemicellulose biomass in terres-trial environments. Various alginate lyases produced by marine microbes have played an important role in the degradation of marine alginate, and several alginate lyases have been isolated from various types of microorganisms over the past few years, especially from brown algae bacteria. Alginate lyases derived from marine bacteria serve as a stable pool of enzymes in the process of alginate degradation and marine carbon utilization. Alginate lyases derived from marine bacteria have great potential for application in the pharmaceutical industry, biofuel production, and environmental protection. It is vital to discover more new alginate lyases and explore their structure, functions, and structure-function relationship in order to advance marine enzymology and biotechnology. Almost no alginate lyase product has been developed for therapeutic applications (such as antibacterial, anticancer, and other diseases). Based on the foregoing review, extensive research in the field of alginate lyases derived from marine bacteria in the direction of advanced biotechnologies is expected.

Conflicts of Interest:
The authors declare that there are no conflict of interest regarding the publication of this paper.