在木糖无机盐培养基中,不同大肠杆菌菌株的生长速率存在较大差异。在缓慢生长菌株Escherichia coli B0013和E.coli B2198中表达快速生长菌株E.coli K12的木糖运输和分解代谢的2个操纵子(xylBA-xylFGH),能够显著提高其利用木糖生长的速率。进一步研究发现,单独表达木糖运输基因xylFGH即可以显著提高菌株起始生长速率。对菌株K12、B0013和B2198的xylFGH基因序列进行测定和分析比对,发现菌株B0013和B2198利用木糖生长缓慢是由于木糖运输载体基因发生无义突变而导致读框中断,B0013的xylH基因中TGG突变为TAG(Trp126*),B2198的xylG基因中CGA突变为TGA(Arg241*),木糖运输效率低下。在B0013中过量表达其自身的另一种木糖运输载体基因xylE或调控基因xylR,可以适当提高其生长速率。但是木糖高效运输系统xylFGH缺陷而导致的生长缓慢表型,并不能够仅仅通过表达低效运输载体基因xylE而显著改善。对菌株B0013的xylH基因进行回复突变,突变株恢复快速生长表型。测序结果发现,xylH基因第126个密码子因无义突变而形成的终止密码TAG,重新回复为Trp密码TGG。

It was found that growth rate of different Escherichia coli strains differed evidently when incubated in xylose minimal medium.Expression of two reversely adjacent xylose metabolism operons(xylBA-xylFGH) from rapid growing E.coli K12 in slow growing E.coli B0013 and B2198 increased their growth rates on xylose.Results showed that expression of xylose transporter gene xylFGH alone also significantly increased initial growth rate of the recombinants.DNA sequencing and analysis of xylFGH in K12,B0013 and B2198 indicated that the 126 thcodon TGG changed to TAG(Trp126*) in xylH of B0013 and the 241 stcodon CGA changed to TGA(Arg241*) in xylG of 2198 which caused interrupted reading frames and therefore deficiency in xylose transporters and was responsible for the slow growth phenotype.Overexpression of its own another xylose transporter gene xylE or regulator gene xylR slightly increased the growth rate of B0013.However,slow growth on xylose due to the deficiency in xylFGH cannot be significantly improved by overexpression of xylE alone because Xyl E is a relatively low-affinity xylose transporter.Reverse mutation of xylH in B0013,verified by DNA sequencing(the 126 thcodon TAG changed back to TGG),restored its rapid growth phenotype.