SSulfoquinovosyl diacylglycerol (SQDG) is a membrane-forming lipid present in photosynthetic organisms as well as in distinct bacteria growing in phosphate-limited environments. Four genes for SQDG biosynthesis were previously identified in Rhodobacter sphaeroides, the operon sqdBDC and sqdA. In this work, we found that SMc02490 of Sinorhizobium meliloti is an SqdA orthologue. Expression of the S. meliloti sqdBDC operon in Escherichia coli results in formation of sulfoquinovosyl glycerol (SQGro), while co-expression of this operon together with smc02490 (sqdA) results in formation of SQDG. Furthermore, SqdA allows for incorporation of exogenous SQGro into SQDG in S. meliloti and in E. coli cultures, suggesting that presence of SqdA in bacteria permit them to use environmental SQGro for the biosynthesis of the membrane lipid SQDG. An in vitro enzymatic assay for the acyltransferase SqdA was developed. Cell-free crude extracts of E. coli expressing sqdA can efficiently convert [35S]-sulfoquinovosyl monoacylglycerol into SQDG using as acyl donor acyl carrier protein. Bioinformatic analyses reveal that this sulfoquinovose acylation pathway for SQDG biosynthesis is delimited to the Hyphomicrobiales (Rhizobiales) and Rhodobacterales orders of Alphaproteobacteria.