Krang from Dimension X could also be a great villain. He might be interested in the human sister because she has some unique knowledge or ability that he wants to use for his own evil plans in taking over Earth. He could send his minions to capture her and the Turtles would have to journey to Dimension X to save her.
It could be Shredder as always. He might see the sister as a new threat or a way to get to the turtles. He could try to kidnap her to lure the turtles into a trap.
Perhaps a new villain emerges. A powerful sorcerer who is angry that the turtles have been turned human by a magic that he believes should be under his control. So he tries to either turn them back into turtles in a more twisted way or simply destroy them in their human forms because he sees them as abominations.
A realistic integration could happen through family ties. Imagine a human character is related to someone who was involved in the accident that created the mutagen. This connection could lead them to the turtles, and they could work together to either reverse the mutagen's effects or use it for good in some way. It also adds an emotional layer to the story as the human has a personal stake in the turtles' world.
Perhaps the sister was created by the same mutagen as the turtles but was separated from them. When she finally finds her brothers, there could be some initial distrust and confusion. But as they get to know each other, she becomes an important part of the team. She might have a different personality compared to the turtles, like being more diplomatic, which could help in dealing with some of the more complex situations they face, such as negotiating with other mutants or humans who are not immediately hostile.
Maybe the human sister is a scientist who is working on a project that could potentially help the Ninja Turtles in their fight against evil. But the bad guys find out about it and try to stop her. Or she could be an artist who uses her art to inspire the Turtles and give them new ideas for their strategies. She might also have a secret past that is somehow connected to the mutagen that created the Turtles.
Leonardo is often popular. In human fanfiction, he might be depicted as a strong and noble leader even in human form, guiding the other turtles and any human allies. He could be the one who tries to uphold justice in a human world.
Perhaps a fanfiction where the human Ninja Turtles travel back in time to the era when the original turtles were created. They could interact with the mutagen that made the turtles what they are and maybe change the course of their own existence in some way. For example, they could try to prevent certain villains from ever being created.
One possible plot could be that the turtles are exposed to a strange mutagenic substance that starts to reverse their mutation and gradually turn them human. They then have to deal with the challenges of living as humans, like going to school and fitting into society while also trying to hide their former turtle identities from those who might not understand.
April O'Neil could also be a main character. She might be the one who discovers the mecha technology or helps the turtles figure out how to use it. Her role as a human ally would be important in a story like this.
April O'Neil is a classic choice. She's a human character who has a long - standing relationship with the turtles. Her skills as a reporter can add an interesting dynamic to the story.