The cyclic-di-GMP diguanylate cyclase CdgA has a role in biofilm formation and exopolysaccharide production in Azospirillum brasilense

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• 作者：Alberto Ram&iacute ; rez-Mata ^{aramirezmata@gmail.com" class="auth_mail" title="E-mail the corresponding author} ; Lilia I. L&oacute ; pez-Lara ^{13_isela13@hotmail.com" class="auth_mail" title="E-mail the corresponding author} ; Ma. Luisa Xiqui-V&aacute ; zquez ^{luisa.xiqui@hotmail.com" class="auth_mail" title="E-mail the corresponding author} ; Sa&uacute ; l Jij&oacute ; n-Moreno ^{jims_pcm@hotmmail.com" class="auth_mail" title="E-mail the corresponding author} ; Angelica Romero-Osorio ^{angy.4behappy@gmail.com" class="auth_mail" title="E-mail the corresponding author} ; Beatriz E. Baca ; ^{beatriz.baca@correo.buap.mx" class="auth_mail" title="E-mail the corresponding author}
• 关键词：Azospirillum brasilense ; Biofilm production ; Cyclic di-GMP ; Diguanylate cyclase activity
• 刊名：Research in Microbiology
• 出版年：2016
• 出版时间：April 2016
• 年：2016
• 卷：167
• 期：3
• 页码：190-201
• 全文大小：2223 K

文摘

In bacteria, proteins containing GGDEF domains are involved in production of the second messenger c-di-GMP. Here we report that the cdgA gene encoding diguanylate cyclase A (CdgA) is involved in biofilm formation and exopolysaccharide (EPS) production in Azospirillum brasilense Sp7. Biofilm quantification using crystal violet staining revealed that inactivation of cdgA decreased biofilm formation. In addition, confocal laser scanning microscopy analysis of green-fluorescent protein-labeled bacteria showed that, during static growth, the biofilms had differential levels of development: bacteria harboring a cdgA mutation exhibited biofilms with considerably reduced thickness compared with those of the wild-type Sp7 strain. Moreover, DNA-specific staining and treatment with DNase I, and epifluorescence studies demonstrated that extracellular DNA and EPS are components of the biofilm matrix in Azospirillum. After expression and purification of the CdgA protein, diguanylate cyclase activity was detected. The enzymatic activity of CdgA-producing cyclic c-di-GMP was determined using GTP as a substrate and flavin adenine dinucleotide (FAD⁺) and Mg²⁺ as cofactors. Together, our results revealed that A. brasilense possesses a functional c-di-GMP biosynthesis pathway.