The biosynthesis of poly(L-alanylglycine) (poly(AG)) was performed in high cell density cultures of recombinant Escherichia coli. The purity of the material was determined by amino acid analysis, elemental analysis, and H-1 NMR spectroscopy. Fed batch fermentation increased the yield of recombinant protein from levels of tens of milligrams per liter (typical of batch fermentation in rich media) to hundreds of milligrams per liter. Poly(AG) comprising 64 diads [(AG)(64)] was recrystallized from dichloroacetic acid solutions in the form of texture-oriented chain-folded lamellae with a lamellar stack periodicity of 3.2 nm. The crystal structure within the lamellar core is similar in general, but different in detail, to the antiparallel beta-sheet structure previously reported for oriented films of poly(AG) and fibers of Bombyx mori silk fibroin (silk II). The structure consists of polar antiparallel (ap) beta-sheets, with repetitive folding through gamma-turns every eighth amino acid (including the fold), stacking with like surfaces together. The wide-angle X-ray diffraction signals index on an orthorhombic unit cell with a (hydrogen bond direction) = 0.948 nm, b (sheet stacking direction) = 0.922 nm, and c (chain direction) = 0.695 nm. The stacking distance (b-value) is increased by about 3% in comparison with the previously reported structure of poly(AG), owing, we believe, to steric interaction at the lamellar fold surfaces. Random shears of approximately +/-a/4 and shears of +/-c/2 in the ac plane are required to obtain a good fit between the calculated and measured X-ray structure factors.