How does one choose an optimal alphabet for finding a Huffman encoding?
The size required to store the Huffman code table scales like the number of codewords. We expect the number of unique $k$-letter words to be exponential in $k$, in fact roughly $2^{kH}$, where $H$ in the source entropy, though since the file is not infinite, for large $k$ we will actually see less. Still, this suggests that for logarithmically large $k$, most of the $k$-letter strings will be almost unique, and so the overall compression for such a $k$ would typically be quite small. In view of that, you can just try several values of $k$, and choose the best one. After you do some such experiments, you can formulate and perhaps prove a hypothesis as to the optimal value of $k$ in different situations.