The angle misalignment error of a USBL (Ultrashort Baseline) acoustic array is one of the major error sources of the strapdown inertial navigation system/USBL positioning system, which will directly affect the positioning accuracy of the USBL positioning system. For the traditional calibration method cannot accurately estimate the angle misalignment error due to its strict trajectory requirements in the field experiment and the high-precision layout of the transceiver array elements, a new method for estimating the angle misalignment error of a USBL acoustic array based on single transponder and dual-vector reconstruction is studied in this paper. The precondition of USBL misalignment calibration is to locate the underwater transponder accurately. In this paper, the single transponder segmentation iterative long baseline method is used to locate the underwater target transponder. The dual-vector reconstruction method is studied to control the estimation accuracy of USBL misalignment error calibration based on the traditional single transponder method, which provides a theoretical basis for the determination of the iteration times to the USBL angle misalignment error estimation module. The underwater experiment results show that the positioning error could be reduced to less than 1 m after the angle misalignment error compensation. The underwater transponder positioning and the angle misalignment error estimation of USBL could be accomplished in a circle sailing. It is a new method with good performance of high estimation accuracy, simple operation, and easy realization.