E is the third note of the C major scale, and mi in fixed-do solfège. It has enharmonic equivalents of F♭ (which is by definition a diatonic semitone above E♭) and D, amongst others.
When calculated in equal temperament with a reference of A above middle C as 440 Hz, the frequency of Middle E (E4) is approximately 329.628 Hz. See pitch (music) for a discussion of historical variations in frequency.
F♭ is a common enharmonic equivalent of E, but is not regarded as the same note. F♭ is commonly found after E♭ in the same measure in pieces where E♭ is in the key signature, in order to represent a diatonic, rather than a chromatic semitone; writing an E♭ with a following E♮ is regarded as a chromatic alteration of one scale degree.