Ghosh Arabinda, Verma Anil Kumar, Tingirikari Jaganmohan Rao, Shukla Rishikesh, Goyal Arun
Department of Biotechnology, Indian Institute of Technology Guwahati, Guwahati, 781 039, Assam, India,
Mol Biotechnol. 2015 Feb;57(2):111-27. doi: 10.1007/s12033-014-9807-4.
Production of manno-oligosaccharides (MOSs) from pretreated and defatted copra meal (dFCO) hydrolysis was achieved by endo-mannanase. Structural characterization of dFCO by FT-IR and NMR exhibited resemblance with galactomannan. The time-dependent hydrolysis of dFCO by recombinant endo-β-(1 → 4)-mannanase of Clostridium thermocellum by TLC and HPAEC displayed the release of mannose and MOSs mannobiose and mannotriose. Purified MOSs yielded 40 % mannobiose and 18 % mannotriose confirmed by mass spectroscopy which showed mannobiose (m/z = 365) and mannotriose (m/z = 527). The homology based structural analysis of catalytic endo-mannanase (CtManT) showed the catalytic core composed of Glu181 and Glu300 acting as acid/base and Glu288 as a nucleophile during galactomannan hydrolysis. Sub-site mapping of CtManT exhibited two aglycone and four glycone sites at cleavage sites existing on either side of β-(1 → 4)-linkage of galactomannan. Isolated MOSs displayed potential prebiotic characteristics and supported higher growth of probiotic Lactobacillus acidophilus and Bifidobacterium infantis than with standard inulin. Moreover, MOSs displayed over 97 % tolerance to simulated gastric juice, intestinal fluid, and α-amylase proving its potential as a stable prebiotic over inulin. In vitro cytotoxicity assay of MOSs (500 µg/mL) on human epithelial colorectal adenocarcinoma cell line (HT-29) demonstrated 60 % decreased viability of cells after 48 h displaying anti-tumorigenic property.
通过内切甘露聚糖酶实现了从预处理的脱脂椰干粕（dFCO）水解产物中生产甘露寡糖（MOSs）。通过傅里叶变换红外光谱（FT-IR）和核磁共振（NMR）对dFCO进行结构表征，结果表明其与半乳甘露聚糖相似。通过薄层色谱（TLC）和高效阴离子交换色谱（HPAEC）对嗜热栖热菌重组内切-β-(1→4)-甘露聚糖酶水解dFCO的时间依赖性分析显示，释放出了甘露糖以及MOSs中的甘露二糖和甘露三糖。通过质谱分析确认，纯化后的MOSs中甘露二糖含量为40%，甘露三糖含量为18%，质谱结果显示甘露二糖（m/z = 365）和甘露三糖（m/z = 527）。基于同源性的催化内切甘露聚糖酶（CtManT）结构分析表明，在半乳甘露聚糖水解过程中，催化核心由作为酸/碱的Glu181和Glu300以及作为亲核试剂的Glu288组成。CtManT的亚位点图谱显示，在半乳甘露聚糖β-(1→4)-连接两侧的切割位点处有两个糖苷配基位点和四个糖基位点。分离得到的MOSs显示出潜在的益生元特性，与标准菊粉相比，能更好地支持益生菌嗜酸乳杆菌和婴儿双歧杆菌的生长。此外，MOSs对模拟胃液、肠液和α-淀粉酶的耐受性超过97%，证明其作为一种比菊粉更稳定的益生元的潜力。对人上皮结肠腺癌细胞系（HT-29）进行的MOSs（500 µg/mL）体外细胞毒性试验表明，48小时后细胞活力降低了60%，显示出抗肿瘤特性。
