Protein folding is essential for the survival of all organisms. Early co- and post-translational folding stages are critical because they determine a protein's ability to function for the remainder of its life. When these early folding steps go awry, proteins form harmful aggregates that cause deadly maladies and render protein production costly in the basic-research, biotechnology, and pharmaceutical settings. However, it is not well understood how proteins avoid aggregation and reach their native state in the cell. In this thesis, fluorescence lifetimes and frequency-domain fluorescence depolarization decays were exploited to determine how proteins attain their functional state within the cellular environment. We first explored co-translational folding. We demonstrate that ribosome-bound globin proteins have a compact region containing only 60-80 amino acids (40-50% of the full-length protein). Remaining residues may interact with the ribosome. We identify chain-length dependent interactions between ribosome-bound proteins and ribosomal proteins, trigger factor, and Hsp70. The size of the compact region demonstrates that at least half the sequence folds post-translationally. Upon release from the ribosome, proteins form either the functional native state or harmful aggregates. These states are kinetically trapped from each other, demonstrating the critical importance of the early folding steps for long-term protein health and function. We show that the Hsp70 molecular chaperone acts during the immediately post-translational folding period to grant solubility to aggregation-prone protein variants. Surprisingly, the soluble fraction contains undesirable soluble aggregates, and higher Hsp70 chaperone concentrations are required to prevent their formation, relative to insoluble aggregates. Finally, much higher than physiologically relevant concentrations of Hsp70 are sometimes required to prevent soluble-aggregate formation. We then identified the Hsp70 binding sites in 2,258 E. coli proteins and found that 99% of these proteins contain at least one binding site. Many Hsp70 client proteins do not require chaperones assistance for solubility, but Hsp70 may enable these proteins to form the native structure. The solvent exposure of Hsp70 binding sites suggests proteins may begin folding while bound to the chaperone. In all, this work provides fundamental insights about cellular function and informs the future development of targeted strategies to overcome deleterious protein-aggregation.