As a half-Austrian half-Spaniard born in Flanders with a life spent both in Germany and Spain, I've always wondered what accent Charles V would have used naturally, in normal conversation. His famous quote, though mis-attributed through a lack of historical evidence, is that he spoke "Spanish to God, Italian to Women, French to Men, and German to [his] Horse", so might it have been more a case that he was so multi-lingual he just used whatever accent came with the language he would have been speaking at the time?
A reply to /u/doc_fontaine
May your fortunes be as bright as Charles', but your genes much more diverse than his! :-D
Charles was raised in Gent and he had limited travel until he came of age and went to Spain to claim that inheritance. He always had a Flemish, Gentois accent. When he first arrived in Spain, he didn't speak Spanish, which led to a negative initial reception by his new subjects. However, he eventually learned Spanish and at the end of his life decided to retire in Spain.
On the other hand, his brother Ferdinand was raised in Spain, spoked Castilian and Catalan, and was well liked by Spaniards. When their father Philip the Handsome died unexpectedly, there were discussions between King Ferdinand and Emperor Maximilian as to how the inheritance should be split. In the end Charles ended up gaining all of it de jure although one of his first decisions as he came to Spain was to order his brother Ferdinand to go to Austria to serve as his regent there. Charles was jealous of Ferdinand's popularity in Spain and worried his subjects there may prefer Ferdinand to be their king.
Charles had a good point however, as a combination of his foreign-ness, his greedy Flemish retinue, and his very heavy demands for funds to secure his HRE emperorship all led to the Revolt of the Comuneros almost as soon as he left Spain for the Low Countries and Germany.
You can read on my profile page that later on Ferdinand became established in Austria and gained the kingship of Hungary for himself. All that set the motions that led to the partition of Charles' inheritance between the Spanish and Austrian Habsburgs.