The investigation involves 430,000 Harley-Davidson motorcycles of model years 2008-2011, with anti-lock braking systems, the NHTSA said on Friday.
"We are aware of the investigation and are cooperating with the NHTSA," Harley-Davidson spokeswoman Maripat Blankenheim said.
The traffic safety regulator said it received 43 complaints, three reports of crashes and two reports of injuries.
The NHTSA, however, added that the riders who made the complaints might have failed to change the brake fluid, which must be replaced within two years.
"While it may be true that the complainants failed to adhere to the...fluid service interval requirement, the consequent sudden and complete loss of brake without warning is a concern," the NHTSA said.
(Reporting by Arunima Banerjee and Ankit Ajmera in Bengaluru; Editing by Kirti Pandey)