MPIase is a glycolipid involved in protein integration in E. coli. Recently, we identified CdsA, a CDP-diacylglycerol (CDP-DAG) synthase, as a biosynthetic enzyme for MPIase. YnbB is a CdsA paralogue with a highly homologous C-terminal half. Under CdsA-depleted conditions, YnbB overproduction restored MPIase expression, but not phospholipid biosynthesis. YnbB complemented the growth defect of the cdsA knockout when Tam41p, a mitochondrial CDP-DAG synthase, was co-expressed, suggesting that YnbB possesses sufficient activity for MPIase biosynthesis, but not for phospholipid biosynthesis. Consistently, a chimera consisting of the CdsA N-terminal half and the YnbB C-terminal half (CdsA-N-YnbB-C) complemented the cdsA knockout by itself, but a chimera consisting of the YnbB N-terminal half and the CdsA C-terminal half (YnbB-N-CdsA-C) required co-expression of Tam41p for the complementation. The biosynthetic rate for CDP-DAG in CdsA and CdsA-N-YnbB-C was much faster than that in YnbB and YnbB-N-CdsA-C, indicating that the N-terminal half of CdsA accelerates CDP-DAG biosynthesis to give the fast cell growth. Therefore, the role of YnbB seems to be as a backup for MPIase biosynthesis, suggesting that YnbB is dedicated to MPIase biosynthesis. A mutant with a high pH-sensitive CdsA8 was unable to grow even under permissive conditions when the ynbB gene was deleted, supporting its auxiliary role in the CdsA function. (C) 2019 Elsevier Inc. All rights reserved.