Triosephosphate isomerase (TPI) is an enzyme that functions in plant energy production, accumulation, and conversion. To understand its function in maize, we characterized a maize TPI mutant, zmtpi4. In comparison to the wild type, zmtpi4 mutants showed altered ear development, reduced kernel weight and starch content, modified starch granule morphology, and altered amylose and amylopectin content. Protein, ATP, and pyruvate contents were reduced, indicating ZmTPI4 was involved in glycolysis. Although subcellular localization confirmed ZmTPI4 as a cytosolic rather than a plastid isoform of TPI, the zmtpi4 mutant showed reduced leaf size and chlorophyll content. Overexpression of ZmTPI4 in Arabidopsis led to enlarged leaves and increased seed weight, suggesting a positive regulatory role of ZmTPI4 in kernel weight and starch content. We conclude that ZmTPI4 functions in maize kernel development, starch synthesis, glycolysis, and photosynthesis.