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Aims l -Fuculose is a valuable rare sugar that is used to treat a variety of ailments, including HIV, cancer, Hepatitis B, human lysosomal disease (fucosidosis), and cardio-protective medications. The enzymatic approach for the production of l -fuculose using l -fucose as a substrate would be an advantageous method with a wide range of industrial applications. The objective of this study is the characterization of recombinant l -fucose isomerase from Paenibacillus rhizosphaerae (Pa-LFI) for the production of l -fuculose from an inexpensive and natural source (fucoidan) as well as its comparison with commercial l -fucose (Sigma-Aldrich). Methods and results Fucoidan, a fucose-containing polysaccharide (FPs), was isolated from Undaria pinnatifida , subsequently hydrolyzed, and characterized before the enzymatic production of l -fuculose. The results elaborate that FPs contain 35.9% of fucose along with other kinds of monosaccharides. The purified Pa-LFI exhibited a single band at 65 kDa and showed it as a hexamer with a native molecular mass of 396 kDa. The highest activity of 104.5 U mg−1 of Pa-LFI was perceived at a temperature of 50°C and pH 6.5 in the presence of 1 mM of Mn2+. The Pa-LFI revealed a melting temperature (T m) of 75°C and a half-life of 12.6 h at 50°C. It exhibited that Pa-LFI with aldose substrate (l -fucose), has a stronger isomerizing activity, disclosing K m, k cat, and k cat/ K m 86.2 mM, 32 831 min−1, and 335 min−1 mM−1, respectively. After reaching equilibrium, Pa-LFI efficiently catalyzed the reaction to convert l -fucose into l -fuculose and the conversion ratios of l -fuculose from 100 g L−1 of FPs and commercial fucose were around 6% (5.6 g L−1) and 30% (30.2 g L−1), respectively. Conclusions According to the findings of the current study, the Pa-LFI will be useful in the manufacturing of l -fuculose using an effective and easy approach that produces no by-products. [ABSTRACT FROM AUTHOR]

L-Fuculose and D-ribulose are kinds of rare sugars used in food, agriculture, and medicine industries. These are pentoses and categorized into the two main groups, aldo pentoses and ketopentoses. There are 8 aldo- and 4 ketopentoses and only fewer are natural, while others are rare sugars found in a very small amount in nature. These sugars have great commercial applications, especially in many kinds of drugs in the medicine industry. The synthesis of these sugars is very expensive, difficult by chemical methods due to its absence in nature, and could not meet industry demands. The pentose izumoring strategy offers a complete enzymatic tactic to link all kinds of pentoses using different enzymes. The enzymatic production of L-fuculose and D-ribulose through L-fucose isomerase (L-FI) and D-arabinose isomerase (D-AI) is the inexpensive and uncomplicated method up till now. Both enzymes have similar kinds of isomerizing mechanisms and each enzyme can catalyze both L-fucose and D-arabinose. In this review article, the enzymatic process of biochemically characterized L-FI & D-AI, their application to produce L-fuculose and D-ribulose and its uses in food, agriculture, and medicine industries are reviewed. • L-FI & D-AI are important L-fuculose, and D-ribulose producing enzymes are reviewed. • All the enzymatic properties of L-FI & D-AI are comprehensively discussed. • Mechanisms for aldose-ketose transformation are discussed. • Application of L-fuculose and D-ribulose is described. • Future prospects are required for further research to industrial uses of L-FI & D-AI. [ABSTRACT FROM AUTHOR]

• L-fucose isomerase was characterized from Caldanaerobius polysaccharolyticus. • Capo-LfIase showed maximum activity at 55 °C with melting temperature T m 82.3 °C and pH 6.5. • The recombinant Capo-LfIase showed 108.2 U mg-1of specific activities towards L-fucose. • It produced 28.12% L-fuculose as the sole product from L-fucose. • Capo-LfIase displayed more than 50% of its original activity after incubation at 55 °C for 20 h. L-fuculose is a rare sugar that is useful for the agriculture and medicine industries. L-fucose isomerase (E.C.5.3.1.25), which is an aldose-ketose isomerase, plays a significant role in producing rare sugars. A recommended L-fucose isomerase gene was cloned from Caldanaerobius polysaccharolyticus and purified with a single band of 65 kDa using nickel-affinity chromatography, with a specific activity of 108.23 U mg−1. The native molecular mass existed with 214 kDa was a trimer. The purified enzyme showed a maximum activity in 1 mM Mn2+ at 55 °C and pH 6.5 with a melting temperature (T m) of 80.3 °C in the presence of one molecule per monomer. L-fucose isomerase from C. polysaccharolyticus (Capo-LfIase) exhibited the highest activity of L-fucose with K m , k cat and K cat / k m values of 94.2 mM, 23854 min−1 and 253.3 min−1 mM−1, respectively. Capo-LfIase showed more than 50% thermostability after 20 h of incubation at 45, 55, 65, 75 and 85 °C. The 9 putative active site residues of the L-fucose substrate were described using a homology model, and the results showed that Tyr440, Met185, Trp499 and Asn527 are the candidates of metal-binding residues, while Ser393, Glu337, Glu302, His528 and Asp361 would be involved in substrate binding. The conversion rate of L-fuculose from L-fucose was almost 28.2%, with 80 g L−1 L-fucose, and no byproduct was found. To the best of our knowledge, Capo-LfIase produces high yield of L-fuculose from L-fucose by enzymatic methods. [ABSTRACT FROM AUTHOR]

• A novel d -arabinose isomerase was characterized from Thermanaeromonas toyohensis. • The enzyme showed activity at 75 °C and pH 9.0 for both d -arabinose and l -fucose. • The specific activities 98 and 85.52 U mg−1 for d -arabinose and l -fucose, respectively. • The enzyme produced 27% (21 g L−1) d -ribulose and 24.88% (19.92 g L−1) l -fuculose. • Its half life at 75 °C was 10 h. d -Ribulose and l -fuculose are potentially valuable rare sugars useful for anticancer and antiviral drugs in the agriculture and medicine industries. These rare sugars are usually produced by chemical methods, which are generally expensive, complicated and do not meet the increasing demands. Furthermore, the isomerization of d -arabinose and l -fucose by D d -arabinose and l -fucose by d -arabinose isomerase from bacterial sources for the production of d -ribulose and l -fuculose have not yet become industrial due to the shortage of biocatalysts, resulting in poor yield and high cost of production. In this study, a thermostable d -ribulose- and l -fuculose producing d -arabinose isomerase from the bacterium Thermanaeromonas toyohensis was characterized. The recombinant d -arabinose isomerase from T. toyohensis (Thto-DaIase) was purified with a single band at 66 kDa using His-trap affinity chromatography. The native enzyme existed as a homotetramer with a molecular weight of 310 kDa, and the specific activities for both d -arabinose and l -fucose were observed to be 98.08 and 85.52 U mg−1, respectively. The thermostable recombinant Thto-DaIase was activated when 1 mM Mn2+ was added to the reactions at an optimum pH of 9.0 at 75 °C and showed approximately 50% activity for both d -arabinose and l -fucose at 75 °C after 10 h. The Michaelis-Menten constant (K m), the turnover number (k cat) and catalytic efficiency (k cat / K m) for d -arabinose/ l -fucose were 111/81.24 mM, 18,466/10,688 min−1, and 166/132 mM−1 min−1, respectively. When the reaction reached to equilibrium, the conversion rates of d -ribulose from d -arabinose and l -fuculose from l -fucose were almost 27% (21 g L−1) and 24.88% (19.92 g L−1) from 80 g L−1 of d -arabinose and l -fucose, respectively. [ABSTRACT FROM AUTHOR]

[Display omitted] • Co-expression of l -fucose isomerase and d -tagatose-3-epimerase was accomplished in E. coli BL21(DE3). • The co-expression system showcased its highest enzymatic activity at 50 °C and pH 6.5. • Recombinant co-expressed enzyme enabled the one-pot production of 6-Deoxy- l -sorbose. • The enzyme produced 16.21 g/L (16.44 %) of 6-Deoxy- l -Sorbose from 100 g/L of l -fucose. 6-Deoxy- l -sorbose (6-DLS) is an imperative rare sugar employed in food, agriculture, pharmaceutical and cosmetic industeries. However, it is a synthetic and very expensive rare sugars, previously synthesized by chemo-enzymatic methods through a long chain of chemical processes. Recently, enzymatic synthesis of rare sugars has attracted a lot of attention due to its advantages over synthetic methods. In this work, a promising approach for the synthesis of 6-DLS from an inexpensive sugar l -fucose was identified. The genes for l -fucose isomerase from Paenibacillus rhizosphaerae (Pr-LFI) and genes for d -tagatose-3-epimerase from Caballeronia fortuita (Cf-DTE) have been used for cloning and co-expression in Escherichia coli , developed a recombinant plasmid harboring pANY1-Pr-LFI/Cf-DTE vector. The recombinant co-expression system exhibited an optimum activity at 50 °C of temperature and pH 6.5 in the presence of Co2+ metal ion which inflated the catalytic activity by 6.8 folds as compared to control group with no metal ions. The recombinant co-expressed system was stable up to more than 50 % relative activity after 12 h and revealed a melting temperature (T m) of 63.38 °C exhibiting half-life of 13.17 h at 50 °C. The co-expression system exhibited, 4.93, 11.41 and 16.21 g/L of 6-DLS production from initial l -fucose concentration of 30, 70 and 100 g/L, which equates to conversion yield of 16.44 %, 16.30 % and 16.21 % respectively. Generally, this study offers a promising strategy for the biological production of 6-DLS from an inexpensive substrate l -fucose in slightly acidic conditions with the aid of co-expression system harboring Pr-LFI and CF-DTE genes. [ABSTRACT FROM AUTHOR]

• L-Ribulose producing l -ribose isomerase was characterized from Cryobacterium sp. N21. • CrL-RIse showed maximum activity at 35 °C and pH 9.0 in glycine-NaOH (50 mmol) buffer. • CrL-RIse showed highest specific activity (54.96 U mg−1) with l -ribose as substrate as compared to other l -RIs. • The recombinant CrL-RIse produced 31 % l -ribulose as the sole product from l -ribose. l -Ribose isomerase (l RI) is an enzyme that can catalyze the reversible isomerization between l -ribose and l -ribulose. It can also perform the conversion between many aldoses into their corresponding ketoses. l -RI was produced from Cryobacterium sp. N21 (CrL-RIse), and l -ribose was utilized as a substrate. The recombinant l -RI gene was cloned and overexpressed from Cryobacterium sp. N21. The purification of CrL-RIse was performed by metal-affinity chromatography. The enzyme displayed a corresponding band with an approximate size of 35 kDa on the SDS-PAGE analysis. The protein for this gene contains 266 amino acids with an expected molecular weight (M w) of 29.6 kDa. The measured M w of CrL-RIse calculated by HPLC was 125 kDa. CrL-RIse was extremely active in glycine buffer at 35 °C, pH 9.0, showing a specific activity of 54.96 U mg−1. CrL-RIse displayed no major increase in activity with metal ions, excluding Mn2+. The estimated Km , K ca t , K cat /Km and V max values of CrL-RIse were 37.8 mM, 10,416 min−1, 275.43 min−1 mM−1, and 250 U mg−1, respectively. The rate of l -ribulose production was 31 % (6.24, 12.11, and 20.89 g L−1) at equilibrium by utilizing 20, 40, and 70 g L−1 of the substrate, respectively. The results indicated that CrL-RIse has the capability to manufacture l -ribulose from l -ribose. [ABSTRACT FROM AUTHOR]

• l -Ribulose producing l -ribose isomerase was characterized from Mycetocola miduiensis. • Mm-LRIse showed maximum activity at 40 °C and pH 7.5 in sodium phosphate (50 mM) buffer. • Mm-LRIse showed the highest specific activity (47.40 U mg−1) with l -ribose as substrate as compared to other l -RIs. • It produced 32 % l -ribulose as the sole product from l -ribose. An enzyme, l -ribose isomerase (l -RI), mostly catalyzes the isomerization of l -ribose and l -ribulose. These so-called rare sugars are essential for the treatment of cancer and other viral diseases. In the present study, l -ribose isomerase produced from a bacterium, Mycetocola miduiensis (Mm-LRIse), by using l -ribose as a carbon source. The recombinant l -ribose isomerase gene was cloned and overexpressed from M. miduiensis and purified with an exclusive band of 32 kDa by nickel-affinity chromatography. This gene possessed 267 amino acids protein having an estimated molecular weight of 29,568.17 Da. The native molecular weight of Mm-LRIse estimated by HPLC was 134.84 kDa. The recombinant l -ribose isomerase was highly active in sodium phosphate (50 mM) buffer at 40 °C and pH 7.5, showing the specific activity up to 47.40 U mg−1. Mm-LRIse showed no significant enhancement in activity with metallic ions except Mn2+ and Co2+. The values of Km , K ca t , K cat /Km and V max of Mm-LRIse against l -ribose substrate were 42.48 mM, 9259.26 min−1, 217.43 min−1 mM−1, and 277.78 U mg−1 respectively. At equilibrium, the l -ribulose transformation rate was nearly 32 % (6.34 g L−1) using 20 g L−1 of l -ribose. The results revealed that the Mm-LRIse enzyme has a potential for L-ribulose production from l -ribose. [ABSTRACT FROM AUTHOR]