Polyhydroxyalkanoates (PHAs) are natural polyesters synthesized by microorganisms under physiological stress. PHAs can act as a potential substituent to petroleum-derived plastic. Pseudomonas sp. is a Gram-negative bacteria and is known for its metabolic adaptability and ability to utilize various carbon sources for PHA production. PHAs have various applications in the fields of environment as well as the medical sector. Pseudomonas sp. are known to synthesize medium chain length PHA (mcl-PHA). These PHAs are biocompatible, biodegradable, and nontoxic. PHAs are synthesized by several metabolic pathways like the tricarboxylic acid (TCA) cycle, fatty acid β oxidation pathway, and de novo biosynthesis. PHA synthase is a crucial enzyme for polymerization of hydroxy-acyl monomers. This chapter focuses on PHA biosynthetic pathways, its regulation, manipulation of carbon catabolite repression, genome-scale reconstruction, and PHA depolymerization with respect to Pseudomonas sp. Owing to its characteristics such as metabolic versatility and mcl-PHA synthesis, Pseudomonas sp. has the potential to be used as a microbial factory for the synthesis of PHA which is advantageous to be used for numerous applications.